September 2020 The Scientist

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| WWW.THE-SCIENTIST.COM | WWW.THE-SCIENTIST.COM SEPTEMBER MONTH 2018 2020

HOW HOMO SAPIENS EVOLVED IN AFRICA SOUTH AMERICAN PREHISTORY UNDERSTANDING MORPHOLOGICAL CONTROL

PLUS

RESEARCHERS’ ROLES IN CRAFTING SCIENCE POLICY

HUMAN PATHS ARCHAEOLOGY AND GENETICS ARE STARTING TO RESOLVE HUMANITY’S ORIGIN AND SPREAD

Designed for one purpose: microbial growth curve analysis

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Announcing The Scientist’s annual Top 10 Innovations Competition Submit your cutting-edge, life-science technology for consideration by a panel of expert judges. The winners will be the subject of a feature article in the December 2020 issue of The Scientist. • An “innovation” is defined as any product that life-science researchers use in a lab: machines, instruments, tools, cell lines, custom-made molecular probes and labels, software, apps, etc. • Products released on or after October 1, 2019 are eligible. • Entries accepted from May 1 to August 21, 2020. For further information, email: [email protected]

SEPTEMBER 2020

Contents

© JULIA GALOTTA; © MARK GARLICK, SCIENCE PHOTO LIBRARY; © N.R. FULLER, SAYO-ART, LLC

THE SCIENTIST

THE-SCIENTIST.COM

VOLUME 34 NUMBER 09

Features

ON THE COVER: ILLUSTRATION BY © JESS SUTTNER

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30

38

New findings from archaeology, genetics, and other disciplines suggest that multiple waves of migrants from North America quickly developed distinctive cultures.

Africa’s sparse fossil record alone cannot reveal the complete story of human origins. Genomic inquiries of both modern and ancient DNA are trying to fill in the gaps.

Understanding biology’s software— the rules that enable great plasticity in how cell collectives build organs and organisms—is key to advancing tissue engineering and regenerative medicine.

BY KATARINA ZIMMER

BY MICHAEL LEVIN

The Peopling of South America

BY SHAWNA WILLIAMS

Becoming Sapiens

The Making of an Organism

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SEPTEMBER 2020

Department Contents 16

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FROM THE EDITOR

Back to School

Many educational institutions, from preschools to universities, are opening this fall in the midst of a global pandemic that threatens much more than our health. BY BOB GRANT

16

NOTEBOOK

Archaeologists detect traces of ancient beer-making; geneticists map complex ancestry patterns in the US population

46 THE LITERATURE A single gene explains the difference between colorful males and muted female red siskin finches; circulating RNA as potential biomarkers of preeclampsia risk; genetic variants linked to microbial diversity in wounds

46

49 SCIENTIST TO WATCH

TUM-WEIHENSTEPHAN; J. HELBING; © KELLY FINAN; DENIS PAISTE, MIT MATERIALS RESEARCH LABORATORY

Ibrahim Cissé: Molecule Watcher BY JEF AKST

52

CAREERS

When Science Meets Policy

COVID-19 has laid bare some of the pitfalls of the relationship between scientific experts and policymakers— but some researchers say there are ways to make it better. BY DIANA KWON

56

READING FRAMES

Confronting Racism in Zoology

A new book explores the history of scientists’ efforts to classify living things. BY DAVID BAINBRIDGE

49

60 FOUNDATIONS Coronavirus Closeup, 1964ANSWER PUZZLE ON PAGE 11 BY ASHLEY YEAGER IN EVERY ISSUE

9 11 58

CONTRIBUTORS SPEAKING OF SCIENCE THE GUIDE

G N I MB L O F L U X J POS E P L S E A L R Y H A L F T I T I S S C H

I S C P U S A T O L L S L T E A AQU A L UNG N R E K I DON N E T S T O B I T T E RN S A W R T I D E A Z OV T N T R U E T R E NCH D Y A

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SEPTEMBER 2020

Online Contents

THIS MONTH AT THE-SCIENTIST.COM: VIDEO

VIDEO

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Shaping Up

The Morphogenetic Code

Amazonian Secrets

See Reading Frames author David Bainbridge of the University of Cambridge discuss how and why women have physiological features different than those of other female animals.

Hear Michael Levin of Tufts University talk about his study of morphogenetics at the Paul G. Allen Frontiers Symposium in late 2019.

Watch researchers travel to a cave deep in the Amazon to search for clues about the first humans to populate the Americas.

AS ALWAYS, FIND BREAKING NEWS EVERY DAY ON OUR WEBSITE.

Coming in the October issue • Crosstalk between the immune system and the nervous system • Progress in identifying neural correlates that help predict human behavior

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Predicting and Overcoming Resistance Using IsoPlexis’ SingleCell Intracellular Proteomic and Metabolomic Analysis Tools

A

big challenge in the field is that cancer cells develop resistance to targeted therapies, and researchers are striving to understand and characterize how these cancer cells are adapting in response to

these therapies. However, the inherent functional heterogeneity in cancer cells makes this challenging. This heterogeneity complicates translating data from genetic profiles or responses into associated functional behaviors or phenotypes. Although cancer cells have been characterized at the genetic and genomic levels, the functional mechanisms impacting protein-driven functional behaviors and activities can only be revealed through singlecell proteomics. IsoPlexis’ single-cell proteomics has helped researchers overcome resistance to targeted inhibitors, leading to better strategies and combination therapies.

Single-cell intracellular proteomics and single-cell metabolomics combine to characterize drug resistance in melanoma cells The constant adaptation of cancer cells poses a great challenge for developing drug-based treatments, as cells that initially respond can quickly adopt drug-resistant states. IsoPlexis’ single-cell proteomics and multiomic technologies can help characterize the mechanisms and underlying

single-cell intracellular proteomic chips provide information on protein-protein

factors driving drug resistance.

interactions and pathway activation or deactivation, emphasizing changes in

In a recent Nature Communications article, James Heath’s team from

protein expression and alterations in phosphorylation profiles.

the California Institute of Technology described using predictive single-cell

The IsoPlexis single-cell proteomics platform, the IsoLight, functions as

intracellular proteomics and metabolomics to identify how a cancer cell line

a proteomic hub that uncovers the true function from cancer and immune

transitions to a final drug-resistant state via two distinct trajectories.1 When

cells. IsoPlexis’ proteomics hub functionally phenotypes adaptive and innate

BRAFV600E mutant melanoma cells are treated with BRAF inhibitors, they

immune cell populations and examines cell populations that shape the

quickly become drug tolerant. To analyze what was happening within these

tumor microenvironment by looking at the secretome, intracellular proteome,

cells to create a drug resistant state, Heath’s team treated these mutant cells

and metabolome. The platform is capable of identifying up to 32 different

with the BRAF inhibitor vemurafenib for varying durations and analyzed them

cytokines and intracellular proteins in a multiplexed manner at the single-

using integrated single-cell intracellular proteomics and metabolomics.

cell level. This is all combined with IsoPlexis’ fully automated plug-and-play

Using IsoPlexis’ proteomic barcoding technology, the researchers

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characterized cellular heterogeneity within cell populations at different

intuitive and publication-ready advanced visualizations to help researchers

timepoints and quantitatively connected multiple timepoints to characterize

further accelerate their therapies.

dynamic heterogeneity on an individual cell level. Cellular state changes became prominent around day 3, as most probed analytes exhibited a sharp

Visit IsoPlexis.com for more information on IsoPlexis’ intracellular proteome

but transitory increase in variance. Indeed, all of the metabolic enzymes

and metabolome technologies.

and signaling phosphoproteins, all metabolic regulators except for one, and all resistant state markers and regulators except for one displayed

References

this phenomenon. This information showed Heath’s team that cancer cell

1. Y. Su et al., “Multi-omic single-cell snapshots reveal multiple independent trajectories to drug tolerance in a melanoma cell line,” Nat Commun, 11:2345, 2020.

responses to a common stimulus may entail multiple divergent functional pathways while still resulting in the same genomic phenotype. Understanding these functional adaptations allowed the team to predict and develop an effective therapeutic combination to overcome this adaptive resistance.

CREDIT LINE

Revealing true function with the IsoLight proteomic hub Proteomic analysis is instrumental in filling the knowledge gap left by genomics, especially at the single-cell level. The IsoLight system from IsoPlexis enables researchers to ascertain true cellular functional phenotypes. Going beyond the limited information provided by cell surface marker expression, IsoCode 01.2018 | THE SCIENTIST

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SEPTEMBER 2020

Contributors Michael Levin spent his childhood, first in Moscow and then in Swampscott, Massachusetts, observing and

musing about the miniature world of insects. Even then, he says, he was amazed that swarms of individuals could share common goals; his interest would later shift to how collections of living cells coordinate the building of complex anatomical structures. Levin enrolled at Tufts University, receiving two bachelor’s degrees, in computer science and biology, before completing a PhD in genetics at Harvard in 1996. His early work focused on how cells organize embryogenesis, and included the discovery of a new “bioelectric language” used by cells to coordinate their activities. Drawing on the parlance of computer science, Levin has been working on modifying these electrical signals—the physiological “software” that dictates body shape in animals—to induce cells to form new structures without making changes to a cell’s genome, what Levin calls the “hardware.” Now, as a developmental biologist at Tufts University, Levin continues to study how decision making in swarms of cells might be exploited for advances in regenerative medicine that could allow researchers to trigger the rebuilding of entire limbs or organs. “If we understand how cells cooperate in these collectives to build complex things, we can ask them to do it again,” Levin says. On page 38, Levin describes recent work from his own lab and others that demonstrates the promise of harnessing biology’s software.

TUFTS PUBLICITY OFFICE; MICHELLE BAINBRIDGE; LINDSEY DEALEY

David Bainbridge admits to having been “weirdly obsessed with how animals work and how we organize them” from a young age, an intellectual quirk he shares with many of the world’s most notable zoologists. Bainbridge received degrees in zoology (1989) and veterinary medicine (1992) from the University of Cambridge before completing a PhD in reproductive biology at the London Zoo’s Institute of Zoology in 1996. Studying the reproduction of red deer (Cervus elaphus) led him to realize how “fascinating and unusual” human reproduction is in the animal kingdom, and his postdoctoral research at the University of Oxford focused on human pregnancy. “Everybody thought that was hilarious, having a vet working in the OB/GYN department,” Bainbridge says. Now, as the university clinical veterinary anatomist at Cambridge, it’s his job to teach young veterinary students the foundations of the anatomy and reproduction of nonhuman animals. In researching the history of his field, what struck him again and again was scientists’ propensity to rank some living things as inherently better than others—animals above plants, for example, and mammals over reptiles and amphibians. He also discovered how this tendency bled into categorizations of humans, with scientists sometimes putting certain racial and ethnic groups above others. “These were not peripheral people; these were mainstream writers and scientists,” Bainbridge says. In “Confronting Racism in Zoology” on page 56, Bainbridge discusses how we can learn from this shameful legacy and acknowledge our prejudices with an eye toward progressive change. Amanda Heidt was undecided on the focus of her study when she started at MiraCosta Community College in San Diego in 2008, so she took both creative writing and marine biology courses. She ended up disliking her English instructor and loving the marine bio class, so she followed the science route. After two years, she transferred to the University of California, Santa Cruz (UCSC), where she majored in marine biology and minored in chemistry. Then, in the master’s program at Moss Landing Marine Laboratories on the Monterey Bay, she worked in the invertebrate molecular ecology lab as a research tech, testing samples from around the world for invasive species. At the same time, Heidt researched meiofauna, tiny organisms that live between grains of sand. “I basically lived in my car for a summer and drove all up and down California, and I sampled all these different beaches,” she says. She brought the samples back to the lab and used metabarcoding to identify what she’d collected, and learned that meiofauna don’t adhere to a strict latitudinal gradient but are found all over. To help support herself through school, Heidt applied for and received a scholarship from California State University, Monterey Bay, and the NPR-member radio station KQED, which happened to come with a summer internship reporting and writing science news at the KQED headquarters in San Francisco. “That was the trial by fire,” she says. And it went well—so much so that it motivated her to wrap up her degree at Moss Landing and enroll in the UCSC science communication master’s program. There, Heidt interned at Inside Science, The Monterey Herald, and Science, before graduating and accepting a position as The Scientist’s summer intern. “I set it as a personal goal . . . to expand the types of stories I write,” she says. With many of the topics she now covers, “it’s not always the most comfortable thing to do, but I have enjoyed that opportunity to learn new things.” 09. 202 0 | T H E S C IE N T IST

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FROM THE EDITOR

Back to School Many educational institutions, from preschools to universities, are opening this fall in the midst of a global pandemic that threatens much more than our health. BY BOB GRANT

10 T H E SC I EN TIST | the-scientist.com

breaks at more than 10 schools. I consider these episodes to be clear cautionary tales. Adding to this anecdotal evidence is the evolving science surrounding the susceptibility of young people. Recent research has combated notions of adolescents being less susceptible to SARSCoV-2 infection, and in fact has suggested that children who contract the virus may carry higher viral loads than adults. To be sure, I am aware of the fact that for children, school is far more than an educational forum. For most, it fulfills vital social needs for peer interaction. And for too many, even for kids within my own district, school offers a respite from abuse, hunger, or an otherwise chaotic or uncertain home life. Taking away that refuge represents something much more dangerous than a delay or alteration in the educational process. But for the millions engaging in virtual learning this semester, I am hopeful that parents, students, and teachers can view the situation in a light that extends beyond the obvious disruption to education as usual. I am hopeful that my own kids and others in their classes will cultivate valuable skills and thought patterns as they navigate the technological and social realities before them. After all, even before the pandemic changed so much about life in 2020, the world was becoming ever more connected, virtual, and off-site. There is a strong probability that the next generation will be learning or working remotely in their lifetimes. So at the start of another school year, one that none of us will soon forget, I commend staff members, teachers, school board members, students, and parents who are braving our new normal, and urge them to see the positive in this difficult situation. g

Editor-in-Chief [email protected]

ANDRZEJ KRAUZE

M

y wife and I, like parents across the US and beyond, are grappling with a very difficult situation at the moment: the start of another school year for our children amidst rampant COVID-19 spread in our state and country. Our local school district has decided that its more than 18,000 students will start the 2020–21 school year with a “remote for all” educational model. Kids from 3 to 18 years old, including our preschooler and two elementary school students, will log in to virtual classrooms to meet with teachers, interact with classmates, and follow a curriculum that had to be reenvisioned in the face of the disruption and forced flexibility wrought by the ongoing pandemic. We followed along intently via livestreamed meetings as our district school board hashed out the details of the coming academic year, hoping that evidence-based thinking and appropriately proportioned risk avoidance would guide its decisions. By and large that was the case, and our board heeded warnings and guidance from state and federal public health agencies. Other schools, including private schools in our area, have chosen to invite teachers, students, and staff members back into classrooms with precautions designed to reduce the risk of SARS-CoV-2 transmission. Frankly, I feel for the cohort that is returning to schools in person this fall. Especially for teachers, who are perennially underappreciated, underpaid, and overworked, being forced back into the classroom seems to add insult to historical injury. In effect, they must choose between their vocation and their health, between the needs of their students and the safety of themselves and their loved ones. My consternation arises from facts on the ground. In Israel, schools across the nation returned to in-person instruction in late May. Within days, one Jerusalem high school reported that 154 students and 26 staff members had tested positive for SARS-CoV-2. Infections rippled into their communities. Eli Waxman, a researcher at the Weizmann Institute of Science and chairman of the team advising Israel’s National Security Council on reopening schools, shared some advice in early August for other countries weighing the benefits and drawbacks of opening schools. “They definitely should not do what we have done,” he told The New York Times. “It was a major failure.” Israel made its decision to reopen at a time when the country was reporting fewer than 100 new infections per day. As of this writing (in mid-August), on the eve of many US schools opening their doors, America is seeing an average of more than 56,000 new cases per day, according to the US Centers for Disease Control and Prevention. Indeed, in Georgia, schools in a suburban district north of Atlanta opened on August 3. By August 12, nearly 1,200 students and staff members had been ordered to quarantine due to COVID-19 out-

beyond the cell cycle

p53 as an Immune System Modulator in Cancer

ISOPLEXIS’ MULTI-OMIC PRODUCTS:

SC Secretome

Functional Phenotype

1

IsoPlexis’ Functional Cytometry

Persistence of adoptively transferred T cells with a kinetically engineered IL-2 receptor agonist Parisi G, et al. Persistence of adoptively transferred T cells with a kinetically engineered IL-2 receptor agonist, Nature Communications, 2020

IsoPlexis’ unique omics allow complete functional phenotyping at single-cell resolution and ultra small sample volume characterization.

1

SC Intracellular Proteome SC Metabolome 2

3

Functional Phenotype Multi-omic single-cell snapshots reveal multiple independent trajectories to drug tolerance in a melanoma cell line Su Y,et al. Multi-omic single-cell snapshots reveal multiple independent trajectories to drug tolerance in a melanoma cell line, Nature Communications, 2020

2 3

CodePlex Secretome

Ultra Small Sample Synergistic IL-6 and IL-8 paracrine signalling pathway infers a strategy to inhibit tumour cell migration

To learn more about IsoPlexis' functional proteomics,

Jayatilaka H, et al. Synergistic IL-6 and IL-8 Paracrine Signalling Pathway Infers a Strategy to Inhibit Tumour Cell Migration., Nature Communications, 8, 15584., 2017

VISIT ISOPLEXIS.COM

For Research Use Only. Not for Use in Diagnostic Procedures.

isoplexis.com

p53 and Immune Effector Cell Interactions 1(1)

p53 activation enhances the expression of Toll-like receptors 3, 8, and 9, thereby promoting pathogen or threat recognition, innate immune cell activation, and host cell apoptosis.2,3

2(2)

p53 activation promotes antigen peptide processing and presentation by upregulating transporter associated with antigen processing (TAP) 1 and 2, as well as endoplasmic reticulum aminopeptidase 1 (ERAP1).4,5 The former moves peptides into the endoplasmic reticulum to facilitate complex formation with MHC-I subunits, while the latter trims peptides for antigen presentation.2 TAP1 upregulation increases the number of antigen-presenting complexes in cancer cells, while ERAP1 upregulation increases the number of peptides available for loading.4

3(3)

Tumor cells upregulate the expression of immune-checkpoint molecules such as PD-L1 to avoid detection by effector T cells. In wild-type cells, p53 activation downregulates surface PD-L1 expression through miR-34 upregulation.6 Because p53 mutations link to excess PD-L1 expression in cancer patients, they may be useful for identifying checkpoint inhibitor therapy candidates.2

4(4)

NK cell

TLR8 CD8+ T cell

p53 pathway modulation varies. For example, p53 activation in carcinoma cells promotes natural killer (NK) cell recognition of damaged or transformed cells by upregulating UL16-binding protein 1 (ULBP1) and ULBP2.7 However, p53 activation in melanoma cells represses ULBP2 translation by inducing miR-34a and miR-34c expression, potentially contributing to immune evasion.8

ULBP2

ULBP1

PD-L1

MHCI

MHC-I Antigen peptides

Immune cell upregulation and activation positively correlates with immunotherapeutic success.9 IsoPlexis' single-cell functional proteomics technology helps researchers identify the cell subsets with high functional activation, the underlying functional mechanisms behind their activation, and the functional effects of their activation.10 In this way, IsoPlexis’ functional proteomics can identify the changes in function that genomics can miss.

ERAP1 3

TAP1/2

4

2 Toll-like receptors miR-34

1

miR-34a miR-34c

TLR3 TLR8 TLR9 p53

p53 and the Tumor Microenvironment (TME) (1)

1

(2) 2

3

Pro-inflammatory mediators in the TME induce epithelialmesenchymal transition (EMT), resulting in cells with enhanced invasiveness and migratory propensity.11 Wild-type p53 counters this effect by degrading the EMT mediator Slug.12

p53 mRNA

Wild-type p53 attenuates inflammation by directly suppressing NF-kB activity.11

(4) 4

Hypoxic conditions such as those found in the TME usually upregulate p53, resulting in cell cycle arrest and increased apoptosis. However, these effects are not present in hypoxiainduced upregulation of mutated or inactivated p53, so TME hypoxia selects for cells with elevated metastatic capabilities and decreased apoptotic susceptibility.14

Slug

IKKβ

NF-KB

1

Chronic inflammation in the TME activates NF-kB signaling, suppressing p53 regulation of cell cycle homeostasis. When combined with an NF-kB-mediated elevation of environmental growth factor secretion, this promotes cell proliferation.13

(3) 3

IκB

Mdm2

Epithelialmesenchymal transition

2 4 HIF-1α

Hypoxia

ROS/NO

Polyfunctionality—the ability for immune cells to secrete more than one cytokine at a time—is linked with improved prognosis in cancer patients. This is especially true for T cells.15 IsoPlexis’ single-cell functional proteomics can define immune cells in the TME, helping researchers understand the mechanics behind senescence, polyfunctionality, and identifying potential candidates for cell-based therapeutics.16

1. 2. 3. 4. 5.

D. Hamroun et al., “The UMD TP53 database and website: Update and revisions,” Hum Mutat, 27:14-20, 2006. J. Blagih et al., “p53, cancer and the immune response,” J Cell Sci, 133(5):jcs237453, 2020. Y. Cui, G. Guo, “Immunomodulatory function of the tumor suppressor p53 in host immune response and the tumor microenvironment,” Int J Mol Sci, 17(11):1942, 2016. B. Wang et al., “p53 increases MHC class I expression by upregulating the endoplasmic reticulum aminopeptidase ERAP1,” Nat Commun, 4:2359, 2013. K. Zhu et al., “p53 induces TAP1 and enhances the transport of MHC class I peptides,” Oncogene, 18:7740-47, 1999.

6. 7. 8. 9. 10.

M.W. Braun, T. Iwakuma, “Regulation of cytotoxic T-cell responses by p53 in cancer,” Transl Cancer Res, 5(6):692-97, 2016. S. Textor et al., “Human NK cells are alerted to induction of p53 in cancer cells by upregulation of the NKG2D ligands ULBP1 and ULBP2,” Cancer Res, 71(18):5998-6009, 2011. A. Heinemann et al., “Tumor suppressive microRNAs miR-34a/c control cancer cell expression of ULBP2, a stress-induced ligand of the natural killer cell receptor NKG2D,” Cancer Res, 72:460-71, 2012. S.D. Saibil, P.S. Ohashi, “Targeting T cell activation in immuno-oncology,” Curr Oncol, 27(Suppl 2):S98-S105, 2020. H. Jayatilaka et al., “Synergistic IL-6 and IL-8 paracrine signalling pathway infers a strategy to inhibit tumour cell migration,” Nat Commun, 8:15584, 2017.

,”

IsoPlexis' Panel List Granzyme B, IFN-γ, MIP-1α, MIP-1β, Perforin, TNF-α, TNF-β, GM-CSF, IL-1β, IL-2, IL-4, IL-5, IL-6, IL-7, IL-8, IL-9, IL-10, IL-12, IL-12-p40, IL-13, IL-15, IL-17, IL-17A, IL-17F, IL-18, IL-21, IL-22, IL-27, sCD137, TGF-α, CCL11, IP-10, RANTES, BCA-1, CXCL1, CXCL13, MCP-1, MCP-4, TGF-β1, sCD40L, Fas, MIF, EGF, PDGF-BB, VEGF, P-PRAS40, P-lkBα, P-NF-kB p65, P-Met, P-p44/42 MAPK, P-S6 Ribosomal, P-Rb, P-p90RSK, P-MEK1/2, P-STAT1, P-STAT3, P-STAT5, P-elF4E, cleaved PARP, alpha tubulin, BCA-1 IsoPlexis has several different kinds of solutions with various panels which can currently detect these cytokines, with even more to come.

beyond the

p53 as an Immune Sys

p53 is arguably most well-known as a cell cy and senescence. Given this, p53 dysfunctio frequently mutated gene in human cancer c cancer pathogenesis goes beyond cell cycl p53 regulates many aspects of the immune s to stimulating paracrine activity. As such, proliferation, but also aid tumo

NK cell 4 TLR8 T cell

How Function Dictates Superh

IL-8 M1 macrophage

IL-6

Polyfunctional cells have su occasionally they secrete m polyfunctional cells predict therapy persistence. Identif functions using single-cell pr development of therapeutics.

2 IFN-γ

Senescenceassociated secretory phenotype

1

These superhero cells play a critical rol solid tumors. Researchers recently used I to understand the functional drivers of T ce solid tumor agonist. Mice treated with the no adoptively transferred cells into tumors. These s response. In a phase 1 clinical trial for melanoma response and clear mechanistic upregulation of p a longer anti-tumor response than traditional AC polyfunctional cells is imperative for creating

Neighbor cells CXL-1 PAT-1 IGFBP-1 3

Angiogenesis

Aberrant polyfunctional cells become super cells contribute to resistance, suppression, o release syndrome. Researchers recently fou to targeted therapies, they lead to increase mutations in GBM tumors along druggable not induced significant improvement for pa universally.21 Measuring and identifying the single-cell proteomics technology is the onl

Thrombospondin-1

p53 and the Secretome (1) 1

The effect of p53 on cytokine signaling can induce senescence in neighboring cells. Cells presenting a p53-induced senescence-associated secretory phenotype (SASP) can induce a variety of functional changes in NK cells, T cells, macrophages, neighboring cells, and can affect blood vessel development.17

(2) 2

SASP cells indirectly promote anti-tumor immune cell phenotypes. Certain cytokines activate NK cells, CD4+ T cells, or macrophages. p53 and NF-kB cooperate to direct macrophage function. Monocytes and macrophages play a critical role in immune suppression.18

(3) 3

SASP cells present elevated expression and production of thrombospondin-1, a potent angiogenesis inhibitor.19

(4)

Immune cells can secrete more than two or three cytokines at a time. Cells that secrete more than two cytokines simultaneously correlate to anti-tumor activity, persistence, durability, and more.

4

11. 12. 13. 14.

I. Uehara, N. Tanaka, “Role of p53 in the regulation of the inflammatory tumor microenvironment and tumor suppression,” Cancers (Basel), 10(7):219, 2018. S.P. Wang et al., “p53 controls cancer cell invasion by inducing the MDM2-mediated degradation of Slug [published correction appears in Nat Cell Biol. 2009 Jul;11(7):914],” Nat Cell Biol, 11(6):694-704, 2009. A.V. Gudkov, E.A. Komarova, “p53 and the carcinogenicity of chronic inflammation,” Cold Spring Harb Perspect Med, 6(11):a026161, 2016. A. Sermeus, C. Michiels, “Reciprocal influence of the p53 and the hypoxic pathways,” Cell Death Dis, 2(5):e164, 2011.

15. 16. 17. 18. 19.

Researchers used IsoPlexis’ single-cell intracellula resistant supervillain cells by analyzing the differe The single-cell analytical approach provided clini combination therapy strategies by identifying tha adaptive mechanism.21

By identifying the highly functional supervillain progression and adverse events, researchers ca functional-omics tools can reveal cells with sup proteome, and metabolome to accelerate thera

J. Rossi et al., “Preinfusion polyfunctional anti-CD19 chimeric antigen receptor T cells are associated with clinical outcomes in NHL,” Blood, 132(8): 804-14, 2018. Y. Xiao et al., “Senescent cells with augmented cytokine production for microvascular bioengineering and tissue repairs,” Adv Biosyst, 3(8):1900089, 2019. E. Pavlakis, T. Stiewe, “p53’s extended reach: the mutant p53 secretome,” Biomolecules, 10(2):307, 2020. A. Lujambio et al., “Non-cell-autonomous tumor suppression by p53,” Cell, 153(2):449-60, 2013. K.M. Dameron et al., “Control of angiogenesis in fibroblasts by p53 regulation of thrombospondin-1,” Science, 265(5178):1582-84, 1994.

The Effects of p53 Mutations

e cell cycle:

Under normal circumstances, p53 signaling prevents excess proliferation, migratio and immune detection. However, mutations to p53 can either abrogate these regu or induce a new set of pro-tumorigenic responses altogether. (1)1

stem Modulator in Cancer

ycle regulator, as well as an initiator of apoptosis on often leads to cancer; in fact, p53 is the most cells.1 However, the relationship between p53 and le dysregulation. Under normal circumstances, system, from facilitating effector cell recognition p53 mutations not only unshackle cancer cell origenesis, growth, and metastasis.

(2)2

(3)3

hero Versus Supervillain Cell Types

uperpowers. Most cells secrete one cytokine, but multiple cytokines simultaneously. These highly unique correlates such as anti-tumor activity and fying and characterizing these cell subsets and roteomics enables researchers to accelerate the

le in developing better immunotherapies for IsoPlexis’ single-cell proteomics methods ell persistence in response to a novel ovel agonist recruited polyfunctional superhero cells predict an anti-tumor patients, the circulating NK cell polyfunctional cell subsets sustained CT treatment with IL-2.20 Identifying immune therapies and vaccines.

rvillains that can lead to disease progression. These or immune-related adverse events, such as cytokine und that when glioblastoma (GBM) cells become resistant ed metastasis. While genomics studies have identified pathways, drugs created to target those pathways have atients, and drug resistance occurs quickly and almost functional adaptations are critically important; IsoPlexis’ ly method available to obtain this information.

ar proteomics technology to overcome ential responses in signaling coordination. ically actionable insights for designing at drug resistance occurred via an

n cells that can lead to disease an help overcome these diseases. IsoPlexis’ perpowers in the secretome, intracellular apeutic development.

20. 21. 22. 23.

G. Parisi et al., "Persistence of adoptively transferred T cells with a kinetically engineered IL-2 receptor agonist," Nat Commun, 11:660, 2020. W. Wei et al., "Single-cell phosphoproteomics resolves adaptive signaling dynamics and informs targeted combination therapy in glioblastoma," Cancer Cell, 29:563-73, 2016. J. Blagih et al., “Cancer-specific loss of p53 leads to a modulation of myeloid and T cell responses,” Cell Rep, 30(2):481-96.e6, 2020. W.A. Yeudall et al., “Gain-of-function mutant p53 upregulates CXC chemokines and enhances cell migration,” Carcinogenesis, 33(2):442-51, 2012.

4 (1)

p53 inactivation in cancer cells promotes recruitment of immunosuppressive CD11b+ cells by secreting MCP1, CXCL1, CXCL10, and M-CSF. This attenuates CD4+ and CD8+ T cell responses. Elevated numbers of Treg cells also exist in tumors.22 Mutant p53 proteins alter cellular secretomes by interacting with other transcription factors. In particular, mutated p53 can modulate NF-kB through mechanisms unrelated to the crosstalk between wild-type p53 and NF-kB. Mutant p53-mediated enhanced NF-kB activity results in elevated expression of the pro-migration chemokines CXCL5, CXCL8, and CXCL12.17,23 Mutant p53 signaling in cancer cells promotes survival and EMT activity through STAT transcription factor pathways, which increases the production of matricellular proteins, key promoters of ECM remodeling, as well as other pro-inflammatory mediators.17,24,25 Mutant p53 promotes angiogenesis by several mechanisms. Several mutant variants upregulate an inhibitor of DNA-binding 4 (ID4), which promotes the expression and secretions of proangiogenic factors such as IL-8 and CXCL1.26 p53 mutations not only eliminate wild-type p53 VEGF suppression upon hypoxia, but can lead to enhanced VEGF synthesis and secretion.27

+ CD11b Treg Tcells cell

MC CX CX M-

Restoring p53 to Fight Cancer Targeting p53 therapeutically originally aimed to inhibit cancer cell proliferation. However, since p53 modulates many immune mechanisms, researchers are now investigating if restoring endogenous p53 function will also restore anti-cancer immune activity.28 (1)

1

Wild-type p53 activation using nutlin-3a induced systemic anti-tumor immunity and tumor regression through a mechanism reliant on p53 activation in both tumor cells and tumorinfiltrating leukocytes. This also elevated numbers of polyfunctional cytotoxic T cells.29

(2)

Some forms of mutant p53 are constitutively more active than wild-type counterparts, offering a potential immune cell recognition target.2 Excess p53 causes B cells to raise autoantibodies against the protein. These have been found in patients with various cancers, indicating potential suitability as a diagnostic biomarker.2,30

(3)

Mutant p53 is immunogenic. T cells in cancer patients specifically target p53 mutations, a phenomenon that has been replicated in the laboratory.31,32 This may offer a potential route to “vaccinate” against cancer cells expressing specific p53 mutations.2,33

2

3

24. 25. 26. 27.

Systemic immunity

Tu regres

R. Schulz-Heddergott et al., “Therapeutic ablation of gain-of-function mutant p53 in colorectal cancer inhibits Stat3-mediat tumor growth and invasion,” Cancer Cell, 34(2):298-314.e7, 2018. G.S. Wong et al., “Periostin cooperates with mutant p53 to mediate invasion through the induction of STAT1 signaling in the esophageal tumor microenvironment,” Oncogenesis, 2(8):e59, 2013. G. Fontemaggi et al., “The execution of the transcriptional axis mutant p53, E2F1 and ID4 promotes tumor neo-angiogenesis Struct Mol Biol, 16(10):1086-93, 2009. A. Narendran et al., “Mutant p53 in bone marrow stromal cells increases VEGF expression and supports leukemia cell growth Hematol, 31(8):693-701, 2003.

IsoPlexis' single-cell intracellular proteome technology has helped researchers overcome adaptive resistance21 and allowed researchers analyzing mutant melanoma cancer cells34 to gain more insight into the transition from drug responsive to drug tolerant. Melanoma cancer cells have demonstrated the ability to quickly become resistant to targeted inhibitors, but with IsoPlexis' technology, the researchers were able to demonstrate that cancer cells don't take just one path to drug resistance. While this makes targeting the resistance mechanism even more difficult, IsoPlexis' technology has been able to identify drug susceptibilities for both pathways to enable the identification of effective combination therapies, providing the necessary data to develop more therapies to combat this drug-resistant cell state.34

on, inflammation, ulatory responses

Cytokine secretion

TLR8 CD8+ T cell

TLR8 CD4+ T cell

Migration

CD11b+ T cell

CP1 XCL1 XCL10 -CSF

s,” Nat

3

IL-8 STAT1 STAT3

NF-kB

E2F1

Inactive p53

Mutant p53

angiogenesis

ID4 4

CXCL1 VEGF

Wild-type p53

3

umor ssion

h,” Exp

Matricellular proteins

2

Epithelialmesenchymal transition Cell survival

1

c y

ted

ECM remodeling

CXCL5 CXCL8 CXCL12

1 2 Polyfunctional CD11b+ cytotoxic T cell T cells

28. 29. 30.

General anti-p53 antibodies

Nutlin-3a Mutant specific T cells

B cells

C. Muñoz-Fontela et al., “Emerging roles of p53 and other tumour-suppressor genes in immune regulation,” Nat Rev Immunol, 16(12):741-50, 2016. G. Guo et al., “Local activation of p53 in the tumor microenvironment overcomes immune suppression and enhances antitumor immunity,” Cancer Res, 77(9):2292-305, 2017. B. Schlichtholz et al., “The immune response to p53 in breast cancer patients is directed against immunodominant epitopes unrelated to the mutational hot spot,” Cancer Res, 52:6380-84, 1992.

31. 32. 33. 34.

S.H. van der Burg et al., “Long lasting p53-specific T cell memory responses in the absence of anti-p53 antibodies in patients with resected primary colorectal cancer,” Eur J Immunol, 31:146-155, 2001. E.V. Fedoseyeva et al., “CD4+ T cell responses to self- and mutated p53 determinants during tumorigenesis in mice,” J Immunol, 164:5641-5651, 2000. A.J. Levine, “p53 and the immune response: 40 years of exploration-a plan for the future,” Int J Mol Sci, 21(2):541, 2020. Y. Su et al., "Multi-omic single-cell snapshots reveal multiple independent trajectories to drug tolerance in a melanoma cell line," Nat Commun, 11:2345, 2020.

beyond the cell cycle

p53 as an Immune System Modulator in Cancer

IsoPlexis is dedicated to accelerating the fight against cancer and a range of the world's toughest diseases with its uniquely correlative, award-winning, cellular proteomics systems. By revealing unique immune biomarkers in small subsets of cells, we are advancing immunotherapies and targeted therapies to a more highly precise and personalized stage. Our integrated systems, named #1 innovation by The Scientist magazine and Fierce, are used globally to advance the field of immune biology and biomarkers as we generate solutions to overcome the challenges of complex diseases.

QUOTES

Speaking of Science 1 6

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—John Moore, a virologist at Weill Cornell Medical College in New York City, expressing doubt about Russia’s recently announced approval of a COVID-19 vaccine without a Phase 3 clinical trial or any scientific publications on the results of early-stage human studies (The New York Times, August 11)

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This is all beyond stupid. Putin doesn’t have a vaccine, he’s just making a political statement.

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Right now, science is being discredited and we have alternative “facts,” including around treatments and what you need to do, so bringing science to Congress is important to maintain science and facts. —Social epidemiologist Natalia Linos, executive director of Harvard’s François-Xavier Bagnoud Center for Health and Human Rights, talking to the Greek newspaper Kathimerini about her run for Congress in Massachusetts’s Fourth Congressional District in the midst of the COVID-19 pandemic (August 11)

© JONNY HAWKINS

BY EMILY COX AND HENRY RATHVON

ACROSS

DOWN

6. Latin for “rainstorm” or “storm cloud” 8. Circular sea formations studied by Darwin 9. Rate of flow per unit area 10. Gear for underwater exploration 11. Thalassic deity 13. Piscatorial equipment 14. Pinniped 15. Heron relatives in reeds and marshes 17. Midpoint between high and low water (2 wds.) 18. Sea on one side of the Crimean Peninsula 20. Aggregate of cells studied in histology 21. Depression over an oceanic subduction zone

1. 2. 3. 4. 5. 7. 12. 15. 16. 19.

Organ of respiration Block, Easter, or Christmas What Titan and Rhea orbit Aquatic invertebrate with tentacles, such as 7-Down Floating collection of algae and protozoa Squarish cnidarian such as a sea wasp (2 wds.) Like Ralph Vaughan Williams’s Riders to the Sea Like a hook with a nightcrawler on it Verne’s __ Thousand Leagues Under the Sea Predator in black and white

Answer key on page 5

09. 2020 | T H E S C IE N T IST 1 1

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DIGITAL PCR AS A UNIVERSAL MOLECULAR DIAGNOSTIC STANDARD Across all areas of life science research, scientists are increasingly aware of the need for the absolute quantification of nucleic acids. To make their analyses actionable, they want to directly measure copies of a molecule, rather than rely on a simple positive or negative result. The challenge, however, is that any tool that quantifies nucleic acids needs to be standardized against a universal reference material produced by an authoritative body such as the World Health Organization. If this practice is not in place, the results obtained using this tool will not be reproducible across different tests, laboratories or locations. Currently, there are no convenient primary reference measurement procedures for quantifying these standards. As a result, it is difficult for research laboratories and diagnostics manufacturers to harmonize their nucleic acid quantification methods and produce reliable assays. Fortunately, several metrology laboratories – labs that study the science of measurement – are seeking to develop a primary reference procedure using digital PCR technology. This technique provides absolute quantification of nucleic acid samples without the need for calibration. Metrology labs hope to prove digital PCR’s worth as a primary reference measurement tool that could be used to quantify reference standards. These standards could, in turn, be used to develop diagnostics and other tests with results that can be traced and trusted.

Reliable RNA and DNA Quantification Developing a universal reference measurement procedure would impact every lab that handles nucleic acids. However, one area where absolute RNA and DNA quantification would be especially useful is in clinical laboratories. To create a reliable in vitro diagnostic, for instance, it needs to be calibrated against a standard that can be traced back to an SI unit. This allows researchers to compare the resulting data directly against different labs to ensure accurate interpretation. Creating a traceable diagnostic starts with a hierarchy of successive calibration steps that ensure each test reports a true quantity based on a certified reference material (CRM) from a national metrology lab. At the top of the hierarchy, metrologists calibrate a primary reference measurement procedure against the CRM. This primary procedure ultimately supports the calibration of a diagnostic test and controls. Each step in this hierarchy introduces uncertainty, so, according to the Consultative Committee for the Amount of Substance (CCQM) of the

International Bureau of Weights and Measures (BIPM), the primary reference measurement procedure must have “the highest metrological properties.”1 It must operate according to a completed, described, and understood protocol, and must have its uncertainty completely written down in terms of SI units. “Any method that’s used as part of a primary reference measurement procedure, at its most accurate, would be SI-traceable,” said Jim Huggett, Ph.D., Principal Scientist (Nucleic Acid Research) in LGC’s Health Science and Innovation Division and Senior Lecturer in Analytical Microbiology at the University of Surrey. Put another way: Its calibration should trace back to a universal standard, or CRM.

Jim Huggett, Ph.D., Principal Scientist in LGC’s Health Science and Innovation Division and Senior Lecturer in Analytical Microbiology at the University of Surrey.

An SI-traceable reference measurement system for nucleic acids would benefit virtually all areas of research and medicine that rely on nucleic acid quantification. In medicine, perhaps the biggest long-term impact will be seen with DNA quantification in cancer. Accurately quantifying tumor load via circulating tumor DNA allows physicians to track both disease progression and treatment response. But for now, the focus of the world and the greatest need for absolute quantification concerns the COVID-19 pandemic. As authorities try to mitigate the spread of the virus, accurate quantification of SARS-CoV-2 viral loads in patients is critical. This information helps authorities track the course of the disease and determine whether someone is contagious, with or without clinical symptoms. In the absence of a reference standard, qPCR is only able to produce a qualitative result (i.e. Ct values). Digital PCR, on the other hand, can fully quantify the virus in copies/μl.2

Variable qPCR Standards In all of these areas, the current go-to method for quantifying primary reference materials is real-time quantitative polymerase chain reaction (qPCR). However, qPCR does not offer absolute quantitation. To interpret a qPCR result in terms of nucleic acid quantity, users need to generate a standard curve using a serial dilution, a procedure that is prone to error and bias. “RT-PCR has played a big role in helping us quantify molecular DNA and RNA, and it’s used extensively,” says Dr. Huggett. “But in reality, it’s actually quite difficult to get a precise measurement and to know the trueness of the result because it’s on a log scale. If you count 500 molecules, is it truly 500 molecules, or is it 50 or 5000? That’s the type of variation you could see.” In fact, digital PCR has already been used to quantify qPCR reference standards, exposing qPCR’s variability and inconsistency. This variability makes qPCR less sensitive to rare genetic variants associated with cancer. It also performs poorly when quantifying minimal residual disease, as seen with low ctDNA concentrations or viral load, further complicating its role as a reliable and reproducible quantification method.

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Another source of qPCR variability stems from differing amplification efficiencies of individual assays. Researchers must interpret the quantity of a target nucleic acid sequence in a sample based on sequence amplification and the subsequently-generated fluorescent levels. This can be an unreliable process, with several factors that can interfere with the results. For example, samples containing highly variable sequences or mismatches between primers or probe sequences, as well as secondary and tertiary nucleic acid structures, can hamper the quantification of the target sequence. qPCR can also be impacted by inhibitors in the sample that reduce the overall level of amplification and affect the final quantification. For qPCR to be accurate, it must be calibrated using a standard curve, such as those produced using a primary reference measurement protocol that involves digital PCR.

Digital PCR as a Primary Reference Measurement Tool

For dPCR, droplets containing individual PCR reactions are generated by a droplet generator. After PCR, droplets are counted by a droplet reader.

Unlike qPCR, digital PCR doesn’t require calibration; rather, it directly quantifies nucleic acid samples, making it eligible as an SI-traceable primary reference measurement procedure. Digital PCR works by directly counting the number of nucleic acid molecules in a sample, bypassing the need for a standard curve, and making it more precise and reliable. The method uses a droplet reader to digitally count the number of target sequence copies in a sample that has been partitioned into tens of thousands of nanoliter-sized droplets. “Count” is a recognized dimensionless SI unit, which means digital PCR could potentially serve as an SI-traceable primary reference measurement procedure for counting DNA copy number concentration. Over the past several years, metrology labs across the globe have demonstrated the accuracy and sensitivity of digital PCR in many areas of research, supporting its potential role as a primary reference measurement procedure. For example, David Dobnik, Ph.D. and his team at the National Institute of Biology, in Ljubljana, Slovenia have even shown that dPCR can accurately and precisely quantify viral titers in plants and genetic modifications in foods.3,4

Samples are prepared in a 96 well plate and then divided into nanoliter-sized droplets, where PCR takes place.

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Along with quantifying primary reference standards, digital PCR is being explored as a tool for evaluating the accuracy and clinical utility of laboratory developed tests (LDTs). The National External Quality Assessment Service (NEQAS) in the UK, for example, uses digital PCR to score LDTs from 1,500 testing labs around the world. Sandi Deans, Ph.D., a member of the UK NEQAS Consortium, believes the biggest impact digital PCR will have as a reference measurement tool in medical testing labs is in cancer. “We do a lot of tumor testing, and tumor DNA tends to be present at low levels,” Deans explained. “We need to make sure we’ve got as much good-quality DNA present as possible to get a reportable result.” Digital PCR helps Deans and her colleagues at NEQAS determine if the samples being used for medical testing contain sufficient quality DNA to produce a true and actionable result. “Detecting these ctDNA molecules means the difference between identifying sequence variants in tumors that can be treated using a personalized approach, or not,” said Deans.

Sandi Deans, Ph.D., Professor of Clinical Genomics at The University of Edinburgh and member of the UK NEQAS Consortium.

What Accurate DNA Quantification Means for the Future of Medicine At its core, a reliable primary reference measurement procedure is the key to many forms of measurement across a diverse range of scientific research. In measuring nucleic acids, accurate calibration ensures that life science researchers and physicians can achieve a result they can trust, publish, and use to make clinical decisions that change lives. With its ability to quantify nucleic acids without the need for calibration, digital PCR fits this need and will likely become an essential tool for laboratories around the world to calibrate their measurements against. We cannot yet predict the full impact of digital PCR’s adoption as a primary reference measurement tool or understand the extent to which the traceability of nucleic acid measurements will affect life science research, but it will undoubtedly bring positive change to clinical molecular laboratories and hospitals around the world.

References 1.

Bunk DM. Reference materials and reference measurement procedures: an overview from a national metrology institute. Clin Biochem Rev. 2007;28(4):131-7.

2.

Liu X, Feng J, Zhang Q, et al. Analytical comparisons of SARS-COV-2 detection by qRT-PCR and ddPCR with multiple primer/probe sets. Emerg Microbes Infect. 2020;9(1):1175-1179.

3.

Mehle N, Dobnik D, Ravnikar M, Pompe novak M. Validated reverse transcription droplet digital PCR serves as a higher order method for absolute quantification of Potato virus Y strains. Anal Bioanal Chem. 2018;410(16):3815-3825.

4.

Dobnik D, Spilsberg B, Bogožalec košir A, Holst-jensen A, Žel J. Multiplex quantification of 12 European Union authorized genetically modified maize lines with droplet digital polymerase chain reaction. Anal Chem. 2015;87(16):8218-26.

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Beer Was Here

F

ive thousand years ago, a pot sat burbling in the corner of a wooden hut perched above Lake Constance in Germany. Researchers such as Marian Berihuete-Azorín, an archaeobotanist at the Catalan Institute of Human Paleoecology and Social Evolution, aren’t sure exactly what was inside the pot, but they have a hunch. At some point, the contents boiled over, leaving behind a scorched, bowl-shaped lump studded with the remnants of ancient malted grains—perhaps early evidence, Berihuete-Azorín says, of beer. The study of beer has long been a focus for archaeobotanists interested in the brew’s cultural and historical impor-

16 T H E SC I EN TIST | the-scientist.com

tance. Some scholars have argued that cereal grains such as barley and wheat may have been used in beer before they were used to make bread. Employers in ancient Mesopotamia paid their workers at least partly in beer rations. “Beer and other kinds of fermented beverages are important in many ancient societies, just like many modern ones,” says Claudia Glatz, an archaeologist at the University of Glasgow. The drink played an important cultural role in forming and cementing social bonds, and iconographic evidence shows that beer was also used in religious and spiritual ceremonies. Beyond that, beer was simply nutritious, providing calories at a time when food could be scarce. Beer, it turns out, is “just all-around goodness,” Glatz says.

SEPTEMBER 2020

BEER VISION: Grains of wheat (pictured)

and barley that underwent malting—one of the first steps in beer-making—show a clear thinning in the walls of their honeycomb-like aleurone cells (yellow).

To track beer through history, archaeobotanists often rely on contextual clues: images and writings that mention beer or equipment from large-scale breweries. The organic evidence, the beer itself, just doesn’t survive. But more and more, scientists are turning to modern techniques to try to recreate ancient brewing conditions, hoping to identify physical or chemical markers of the beer-making process that could aid the study of archaeological remains such as grains or microscopic residues embedded in ancient vessels.

TUM-WEIHENSTEPHAN; J. HELBING

Notebook

dried and charred each batch of barley in an oven to simulate the processes that create the charred lumps found at archaeological sites. After “countless experiments,” she was left to make sense of hundreds of photos and microscopy images of blackened grains. The hope, she says, was that they might reveal some consistent feature that would link them to ACOs from the field.

Beer and other kinds of fermented beverages are important in many ancient societies, just like many modern ones. —Claudia Glatz, University of Glasgow

It would take a group workshop with almost two dozen collaborators to prompt the “Eureka” moment. As the researchers pored over the images

together, a predictable pattern finally emerged: the outer layer of plant cells, or aleurone cells, in the treated grains showed unusually thin cell walls— a result of the malting process, BerihueteAzorín says. The team’s results, published this May in PLOS ONE, offer a new way to probe the archaeological record for beer (15:e0231696, 2020). “We went to the microstructure, and it turned out to be a really promising direction to develop,” says Elena MarinovaWolff, an archaeobotanist at the BadenWürttemberg State Office for Cultural Heritage and a coauthor on the new study. “These alterations can be diagnostic, and they lead us to beer.” Drawing on their access to archaeological sites around the world, the researchers went looking for their marker in archaeological samples. They began with ACOs pulled from sites in Egypt that dated to the fourth millennium BCE and have a wellcorroborated history of beer brewing. Both Hierakonpolis in Upper Egypt and Tell elFarkha, a site 120 kilometers northeast of

ANDRZEJ KRAUZE

This is exactly what BerihueteAzorín and her colleagues were aiming to do when they came together a few years ago to study the culinary practices of early Europeans. While excavating ancient sites, members of the team sometimes came across blackened, carbonized lumps—also known as amorphous charred objects (ACOs)— containing cells from cereal grains. These cells are circumstantial evidence, Berihuete-Azorín says, of food preparations that could include beer brewing, although they are rarely studied due to their rarity and the difficult nature of analyzing them. “You can study hundreds of these lumps and find only a few cells,” she tells The Scientist. Berihuete-Azorín needed to simulate archaeological preservation through charring to know which morphological features of grain remained consistent. She collected samples of modern barley grains that had been malted—soaked in water until they began sprouting—at different stages of germination. She then

09. 202 0 | T H E S C IE N T IST 1 7

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Cairo, contain massive vats and other tools associated with large breweries. ACOs retrieved from these areas contained cells with the expected thinning of the cell walls. Seeing that, “we were sure that people prepared the mashes and malted the grains,” Marinova-Wolff says. “It was an amazing feeling.” They next tried samples from three sites in Europe where archaeological evidence for beer-making is less clear. The HornstaadHörnle and Sipplingen-Osthafen sites were once home to Neolithic villages on the shores of Lake Constance, while a third site, excavated while building a car park for the Parkhaus Opera in Zurich, housed a similar

BURNED UP: Researchers suspect that this

authors as “waterlogged,” a potential source of doubt for the team’s marker. In addition, cell wall degradation only points to malting—a step before the final fermentation process needed to create beer. Glatz, in her work studying beer consumption in Mesopotamia, has developed chemical markers using the residues of ancient beer drawn from ceramics. Employing gas chromatography, she identified a set of chemicals that appear both in modern fermented beers and in those ancient residues. A combined analysis of grain cell walls and chemical residues, she says, could form a complementary set of tools that cover the entire beer-making process. “Any method that adds another avenue to try and understand a particular archaeological question is a great one,” Glatz says. “That’s exactly what this paper does, it adds something else into our toolkit.” —Amanda Heidt

ÖAW-ÖAI; N. GAIL

amorphous charred object was formed after the contents of a pot boiled and burned from the outside in.

settlement in Switzerland. All date to around the same time as the Egyptian sites but lack the obvious brewing infrastructure. However, several ACOs taken from these sites showed the same thinning in the cell walls that the team had seen at the other sites, offering strong circumstantial evidence for what would be the oldest known example of malting in central Europe. Ben Schulz, a biochemist at the University of Queensland who studies how proteins change in beer during modern brewing, says he found the team’s methods “interesting and generally quite robust,” but urges further validation. It’s possible that barley could just have gotten wet—something that would trigger spontaneous germination—without any human involvement, and that that would “look very similar to what would happen” during malting, Schulz says. While Egypt is dry and arid, the European ruins were described by the

18 T H E SC I ENTIST | the-scientist.com

Geotyping When Chengzhen Dai set out to investigate the influence of US geography on human genetics a few years ago, the study made a somewhat unusual addition to the work of MIT’s SENSEable City Lab, whose projects typically focus on solar power, climate, waste streams, and other urban questions. But Dai saw it as fitting. Researchers at the lab, where he was doing his master’s, are interested in how humans move around and interact with one another, he explains. As he and his colleagues planned the study, “we had the hypothesis that cities, and in a broader sense, geography has played a major role in how ancestry and admixture occurs.” Dai, now a software engineer at the Institute for Systems Biology in Seattle, and his advisor, designer and engineer Carlo Ratti, teamed up with population

geneticist Alicia Martin of the Broad Institute and other colleagues to test their hypothesis using data from National Geographic’s Genographic Project, a now-discontinued effort to sequence genomes from around the world to track migration patterns. Focusing on the genomes of 32,589 participants who’d provided a postal code, the team compared single-nucleotide polymorphisms (SNPs) in those genomes with the SNPs present in ref-

Better knowledge of human genetic diversity and patterns of admixture is important not only for understanding individuals’ origins, but also for harnessing genetic findings in clinical practice.

erence genomes curated by the 1000 Genomes Project, a UK-based initiative that compiled genomic data on 2,504 people representing 26 populations worldwide. Those SNP data revealed substantial diversity of ancestral origins for each demographic group. For example, Hispanics and Latinos tended to have a mixture of African, European, and Native American ancestry, but the makeup and proportions of these ancestries varied widely among individuals. The researchers also looked for unbroken chunks of the genome, known as haplotypes, that were shared among two or more members of the Genographic cohort. This approach can reveal when two people share a common ancestor within the past 10 to 15 generations, says Gillian Belbin, a population geneticist at Mount Sinai Hospital’s Institute for Genomic Health who uses approaches that explore relat-

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NOTEBOOK

edness based on these segments but was not involved in the new study. It’s similar to what commercial testing companies use to point their customers toward distant relatives in their databases. At a population level, discerning relatedness in this way can reveal the effects of factors such as migrations and admixture that have shaped a group over recent history, she adds. As Dai and his colleagues had predicted, genetic relatedness turned out to correlate with geography, although the drivers of that relationship seemed to vary among demographic groups. Among African-Americans, for example, there was broad genetic relatedness among people living along the East Coast, from Florida to Maine, indicating frequent migration within that area. But the researchers also found evidence of reduced migration through certain regions. Overall, the authors write, the patterns they found are consistent with movement trends during the Great Migration of the early- to mid-20th century, in which millions of AfricanAmericans left the South for cities in other areas of the country. Two other demographic groups the researchers analyzed—European-Americans, and Hispanics and Latinos—had their own distinctive patterns of migration. Another geographic finding was of five distinct clusters of related people within the Hispanics/Latinos category, each of which tended to live in particular areas in the lower 48 states based on the postal codes they’d reported. Only one of these showed strong ancestral links to a place outside the continental US: A cluster made up mostly of people living in central Florida and the New York City area reported that most of their grandparents had been born in Puerto Rico. People in another of the clusters, who reported having grandparents born in the US, Mexico, 20 T H E SC I EN TIST | the-scientist.com

 A

 B

 C

and Cuba, among other countries, tended to live in southern Florida and parts of Texas and California. A third, containing people whose grandparents were predominantly born in the United States, lives mainly in New Mexico and Colorado, as well as parts of California.

The findings of multiple clusters within the category of Hispanics/Latinos, Belbin says, “indicates that there’s some interesting gene flow and population structure perhaps . . . that is not being effectively captured by the self-reporting labels [for race or other

AM J HUM GENET, 106:371–88, 2020

STOP AND GO: Regions of higher- (blue) and lower-frequency (brown) migration, as inferred from the genomes of African-Americans (A), Hispanics and Latinos (B), and European-Americans (C).

We had the hypothesis that cities, and in a broader sense, geography has played a major role in how ancestry and admixture occurs. —Chengzhen Dai, Institute for Systems Biology, Seattle

demographic categories] that are available.” In her studies on Mount Sinai’s patient biobank, Belbin says she’s similarly found clusters of genetically related people that aren’t captured by existing demographic labels. While this is far from the first study of genetic diversity in the US, it’s exceptionally comprehensive, says Scott Williams, a population geneticist at Case Western Reserve University in Cleveland who was not involved in the work. “It’s quite amazing how much information was in there, and the analyses were really complete,” he says. For example, he notes, the research-

ers included people of East and South Asian descent living in the US, who have been left out of most studies of genetic variation and distribution. The team found that people of East and South Asian descent in the US were on the whole less admixed than other groups, likely because they or their ancestors tend to be relatively recent arrivals in the country, says Dai. “In the United States, immigration wasn’t really amenable to [East] Asians and South Asians” until the mid-1960s, he explains. As a result of that lack of mixing, the study found, people of Asian

descent in the study formed well-defined genetic clusters that roughly correlated with their ancestral countries of origin. “These are very . . . distinct populations, and they’re very diverse,” Dai says. “But a lot of times, genetic studies . . . group them as just like one continental ancestry,” mainly because of small sample sizes. This continent-level grouping is “not a very accurate way of reflecting and capturing the genetic diversity of these individuals,” he adds. Better knowledge of human genetic diversity and patterns of admixture is important not only for understanding individuals’ origins, but also for harnessing genetic findings in clinical practice, Williams notes. “Until we know about these patterns of diversity, it becomes very difficult to translate genetic findings for risk of disease easily across populations.” —Shawna Williams

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The Peopling of South America

New findings from archaeology, genetics, and other disciplines suggest that multiple waves of migrants arrived from North America and quickly developed distinctive cultures.

I

n 2007, while searching for signs of ancient human inhabitants of the Andes mountain range, more than 4,300 meters above sea level, Kurt Rademaker came across a field littered with chunks of obsidian—some of them fashioned into tools. “There were hundreds and hundreds of them,” recalls Rademaker, then a graduate student at the University of Maine. “Adjacent to this open-air workshop, just up above it on the hillside, was a beautiful rock shelter. . . . I just had a gut feeling that it was the kind [of site] I had been looking for.” A decade earlier, Rademaker’s advisor, archaeologist Daniel Sandweiss, had made the unexpected discovery of flakes and tools made of obsidian, a volcanic rock, in excavations at one of the oldest archaeological sites in South America. Quebrada Jaguay, which dates to 13,000–11,000 years ago, sits along the Peruvian coast, where there are no natural deposits of obsidian. Sandweiss and his colleagues had analyzed the elements in the artifacts and found that they were likely from a source in the nearby Andes. The researchers suggested that ancient people at Quebrada Jaguay had probably split their time between the coast and the highlands.1 Curious about the early inhabitants of the Andes, Rademaker used

22 T H E SC I EN TIST | the-scientist.com

the obsidian finding as a clue and began searching for early highland sites in an area with deposits of the rock. Rademaker struck pay dirt with his 2007 discovery: the site had clear evidence of human activity, including beads made of bone and hundreds of stone tools. Carbon dating of food residue and collagen from animal bones on the floor of the rock shelter, dubbed Cuncaicha, pinned that activity to as early as 12,500 years before present (BP), more than 1,000 years before the oldest previously dated Andean archaeological sites.2 The finding of such early occupation of the Andes upended some assumptions about the ancient people of the Americas, says Rademaker, now a professor at Michigan State University. For example, scientists generally thought that before 11,000 or so years ago, the Andes would have been covered with glaciers or otherwise too cold for humans to inhabit. But in addition to their archaeological work on the site, analyses of the geology at Cuncaicha revealed that “ice was already well in retreat way before people arrived,” Rademaker says, and the environment would have been about as warm as it is today, temperate enough for human habitation.

CREDIT © SHUTTERSTOCK.COM, LINE DICOGM

BY SHAWNA WILLIAMS

Researchers had also previously thought that the low-oxygen environment of the high Andes would have forced ancient people to stick to lower altitudes after their initial migration into the continent, says Rademaker. Yet Quebrada Jaguay and other nearby sites along the coast have been dated to about the same time period as Cuncaicha. “That suggested a more rapid colonization process of a very challenging environment,” he says. Research on Cuncaicha is just one of the many recent inquiries into the peopling of South America that are challenging previous assumptions about how migrations, cultures, and ancestries shaped that continent’s human landscape. In addition to archaeological clues, genetic investigations of ancient and modern humans are yielding intriguing new details about when and how the continent was populated. But many fundamental questions remain unresolved, such as when people first arrived in South America and how they spread across the continent, and a coherent picture of its prehistory has yet to emerge.

A shifting archaeological picture When anthropologist Tom Dillehay, now at Vanderbilt University, began working at a site called Monte Verde near the southern tip of Chile in 1977, most archaeologists thought the first humans moved into South America from North America about 11,000 years ago, he says. But in 1988, he and a colleague published an analysis of traces of human occupation at Monte Verde—including stone tools, mastodon bones with cut marks, and hearths with traces of burned plants—that were dated to 12,500 BP.3 (They also unearthed charcoal and what might have been stone tools from a layer dated to 33,000 BP, but Dillehay now says he doesn’t think the evidence is strong enough to support occupation of the site that early.) More recently, Dillehay’s group reported new excavations that unearthed human-linked materials dated to between 18,000 and 15,000 BP.4 That timeline positioned Monte Verde as the oldest archaeological site in South America, says Lars Fehren-Schmitz, a biological anthropologist at the University of California, Santa Cruz, who was not involved in Dillehay’s work. The site “shows that people must have reached southern South America quite quickly after they even entered North America.” While the timing is unclear of humans’ first migrations out of a region known as Beringia—which spanned parts of present-day Siberia and Alaska—and into North America, most archaeologists put the date at no more than about 20,000 years ago. The dating of Monte Verde suggests, then, that people made it from one end of the New World to the other—about 10,000 miles—remarkably quickly. Several other sites in South America similarly point to a quick dispersal, Fehren-Schmitz notes. “We actually have a bunch of archaeological sites that are securely dated that fall into the time range of 14,000 to 12,000 BP, that tell us that people were more or less all over South America quite early after the initial peopling of the Americas in general.” There’s wide variation in the style of artifacts found at different early sites, such as projectile points and stone tools used for cutting and scraping, indicating that groups on the continent quickly developed diverse cultures and technologies, Dillehay 24 T H E SC I EN TIST | the-scientist.com

says. “People adapted to a very wide range of different environmental zones and habitats,” exploiting the various food sources they offered. Some studies point to an even earlier occupation of South America than that indicated by Dillehay’s most recent work at Monte Verde. For example, research on a site in Brazil yielded stone artifacts associated with charcoal remnants, suggesting the possibility of cooking fires, that researchers carbon dated to between 35,000 and 28,000 BP.5 And a site in Uruguay called Arroyo del Vizcaíno, dated to between 30,000 and 27,000 BP, yielded more than 1,000 bones of ancient animals, some with apparent cut marks.6 It’s not widely accepted that such remnants truly represent evidence of ancient human activity. In some cases, what are presented as artifacts likely aren’t really human-made, says Dillehay; other times, genuine artifacts might have been carried by floodwaters into layers that don’t reflect their true ages. Eric Boëda, an archaeologist at Paris University Nanterre, says the lack of consensus that humans occupied the Americas before the period known as the Last Glacial Maximum, which extended from about 26,500 to 20,000 years ago, reflects “denialism” in the field. Boëda, who has worked at archaeological digs all over the world, including some of the Brazilian sites with purported human activity dating to more than 20,000 years ago, says artifacts that would be quickly accepted as human-made tools if they turned up in Europe or Africa are discounted when they are discovered in Brazil because of a longstanding belief that the Americas weren’t home to any people until more recently.

In late 2018, two studies compared human genomes from across the Americas to infer that there were multiple waves of migration from North to South America.

Boëda says there’s not enough evidence yet to understand how and when people first reached South America or what subsequent migrations might have taken place to or within the continent. One possibility, he says, is that people came from East Asia through Alaska via an ice-free corridor more than 24,000 years ago, before complete glaciation made overland travel through North America impossible during the Last Glacial Maximum. (See “Where Early South American Migrants Came From” on page 29.) In the view of André Strauss, an archaeologist at the University of São Paulo in Brazil, such an early occupation is possible—but he says he thinks that such an extraordinary claim requires more evidence. “I don’t find any of the evidence available at this point [convincing] without a doubt,” he writes in an email to The Scientist. One gaping hole in the evidence is the lack of human remains in the Americas from before the Last Glacial Maximum, he adds— “not a single tooth!”

KURT RADEMAKER; ALBERTO BARIONI; ANDRÉ STRAUSS

Indeed, the oldest human remains found in South America thus far date to just 10,000 years ago or so. But while these fossils don’t shed much light on how long the continent has been occupied, they can reveal a lot about its ancient inhabitants. For example, skeletons at the Brazilian cave Lapa do Santo are among the earliest remains found in South America, and they paint an intriguing picture of the people who called it home, says Strauss, who’s worked extensively on the site. As he and his colleagues reported in 2016, some skeletons at Lapa do Santo revealed signs of elaborate mortuary rituals involving cutting apart bodies, burning parts of them, and burying several individuals together. And while archaeologists have long envisioned early South Americans as highly mobile hunter-gatherers, he says, isotopic analyses revealed that the people at Lapa do Santo consumed food and water from a circumscribed area that he estimates to be around 1,000 square kilometers, and were thus relatively settled.7 As archaeologists continue to search for more human remains to help flesh out the continent’s prehistory, researchers are also mining modern genomes, together with ancient DNA from sites such as Lapa do Santo, for data that reveal more about ancient South Americans.

Clues in the genome In late 2018, two studies compared human genomes from across the Americas to infer that there were multiple waves of migration from North to South America. One of the studies, led by geneticist David Reich of Harvard University, analyzed both ancient and modern genomes and surmised that before European settlers arrived, there were four waves of migration into South America. One wave was made up of people genetically related to the Clovis culture, distinguished by a style of tools found across much of North America, the researchers concluded. Beginning around 9,000 years ago, that lineage was partly replaced by a separate wave of migrants to South America.8 The second study, led by Eske Willerslev of the University of Copenhagen, included 15 ancient genomes unearthed in North and

DIGGING DEEP: Ancient sites such as the Cuncaicha rock shelter (left) in

Peru and Lapa do Santo in Brazil, where archaeologists unearthed this skull (right, top) and projectile point (right, bottom), are adding new details to the picture of South America’s prehistory.

South America. It found evidence for two waves of migration into the southern continent, one about 14,000 BP from North America, followed by a dispersal of people who had diverged from North American groups and spent time in Central America.9 While the results are somewhat similar to those of the paper from Reich and colleagues, Willerslev says his study didn’t find the widespread replacement of the first migrants that Reich’s did. Rather, Willerslev’s data suggested the two groups intermixed with one another. Willerslev’s team also turned up a puzzling signal in 10,000-yearold skeletons from Brazil: between 2 percent and 6 percent of their genomes were more closely related to modern-day inhabitants of the Andaman Islands in Southeast Asia than to any other population. Willerslev points out that the ancestors of the Andaman Islanders once occupied a much broader swath of East Asia, making it plausible that some could have crossed the land bridge into North America—although that leaves open the question of why their genes haven’t appeared in any ancient or modern North American samples. As for the possibility that they could have crossed the Pacific by boat, “in many people’s minds, at least, it’s a very unrealistic scenario . . . to have done a journey like that” so early in prehistory, Willerslev says. How genetic traces of this population have shown up in Brazil and nowhere else in the Americas “is still a mystery,” he adds. Other recent genetic studies have homed in on what occurred in particular regions during South America’s prehistory. In a study published shortly before the Reich and Willerslev papers, for example, a team led by Anna Di Rienzo of the University of Chicago compared the whole-genome sequences of ancient human remains found in the Andes with the sequences of modern people living in the Andes 09. 202 0 | T H E S C IE N T IST 2 5

Site: Rock shelter at 4,480 meters in elevation dated to ~12,400 BP Contains: Remains of plants and animals consumed as food and other human-made debris; human remains; stone tools Significance: Oldest known site in Andes; shows humans had adapted to high altitudes

CUNCAICHA

QUEBRADA JAGUAY

Site: Remains of a seasonally occupied fishing village dated to ~13,000–11,000 BP Contains: Seafood remnants; hearths; tools made of obsidian and other types of stone Significance: Demonstrates that people were using marine food sources and, together with Cuncaicha, that coastal people had contact with the Andean highlands

ARROYO DEL VIZCAÍNO*

EVIDENCE OF EARLY HUMAN PRESENCE IN SOUTH AMERICA

Excavations of South American sites containing traces of ancient human activity have suggested that humans reached the southern region of the continent at least 14,500 years before present (BP)—remarkably quickly after first entering the Americas—and that they soon developed diverse technologies across different sites. But the picture yielded by these archaeological investigations is a patchwork, leaving open key questions, such as whether the first humans migrated south along the Pacific coast or by some other route. The history is further complicated by disputed claims (examples marked by red headers with asterisks) that certain sites reflect a much earlier occupation of the continent beginning more than 20,000 BP. 26 T H E SC I EN TIST | the-scientist.com

Site: Settlement dated to ~18,500– 15,000 BP Contains: Remains of plants and animals consumed as food; charcoal; wooden artifacts; stone tools Significance: Pushed back the date of earliest known human occupation of the Americas by as much as 5,000 years

© JULIA GALOTTA

MONTE VERDE

TOCA DO SÍTIO DO MEIO*

Site: Rock shelter with signs of human occupation dated to ~35,000–28,000 BP Contains: Charcoal remnants; purported stone tools Criticisms include: Rocks resembling stone tools could have come about through natural geological processes or been made by monkeys.

LAPA DO SANTO

Site: Cave with signs of human activity dated to as early as ~12,700–11,700 BP Contains: Remains of 50 people, dating as far back as 10,600–9,700 BP, who were buried at the site; stone tools; rock art; animal remnants Significance: Yielded ancient DNA for analysis and new insight into early cultures

Site: Assembly of more than 1,000 animal bones dated to ~30,000– 27,000 BP Contains: Bones of giant sloths and other large animals, some with apparent cut marks that may indicate they were butchered by humans; purported stone tools Criticisms include: The bones could have been carried to the site by flowing water rather than human activity; the scenario the authors propose (including human transport of large, killed animals) is not consistent with the way known hunter-gatherer groups operate.

and nearby lowlands. While that study didn’t indicate when people might first have begun settling the Andes, it did reveal that the highland and lowland populations began to diverge around 9,000 years ago, says coauthor John Lindo, an anthropologist now at Emory University.10 “That’s pretty early to start [having established populations] at 9,000 years, I think,” he says. “That’s pretty amazing.” Fehren-Schmitz agrees, noting that in contrast to Eurasia, which experienced multiple mass migrations of humans, South American prehistory overall shows a pattern of populations staying relatively stationary after the initial peopling. In the case of the Andes and nearby coastal regions, he says, “the [west] coast is actually only a very narrow stretch of land and [then] the Andes directly start, so we’re talking about distances between populations of a day or two days of travel— like fifty to one hundred miles. And still we see patterns of genetic distinctness between these groups, which is quite stunning.” Based on current evidence, says Tábita Hünemeier, a population and evolutionary geneticist at the University of São Paulo in Brazil who conducts genetic studies of indigenous Brazilian groups, presentday Andeans seem to have descended from the wave of migrants that replaced the Clovis-related population, together with a more recent wave that occurred around 4,000 BP. After the highland and lowland populations divided around 9,000 years ago, the highland population further split into northern and southern populations around 5,800 BP, a study led by Reich and Fehren-Schmitz recently suggested.11 Less is known about the dynamics of ancient Amazonian groups, says Hünemeier, but genetic evidence points to an initial arrival of people in the first large migration event. That population appears to have been replaced by a second wave of migration into the continent, and linguistic evidence indicates these people quickly differentiated into different groups beginning around 9,000 years ago, leading to a panoply of languages, more than 100 of which still exist today. The current goal of her work, Hünemeier says, is “to figure out how [people in the second group] settled Amazonia and how they split,” as well as “which was the original group that came to Amazonia.” In a study published earlier this year, she and her colleagues found modern genetic traces of a population known as the Tupí that dominated the Brazilian coast during the 15th century but had supposedly been driven extinct by European conquerors.12 Modern people known as the Tupiniquim, who live in cities and do not speak an indigenous language, have long identified themselves as descendants of the Tupí, and the researchers found that they do indeed have signals of indigenous ancestry, along with European and African heritage, in their genomes. Furthermore, their indigenous ancestry was distinct from that of any other modern groups who have been studied. The team’s analysis points to a migration of Tupí people from central Amazonia northeast toward Brazil’s coast around 2,000– 3,000 years ago, followed by a later migration of a separate Tupí group who traveled southeast but also ultimately expanded along the coast. They were not the first people to reach the Brazilian coast; archaeological sites with distinctive shell mounds known as sambaquis indicate that others had settled the coast 10,000–8,000 years ago, but they appear to have been completely replaced by the Tupí. “We don’t know where [the people who made the shell 09. 202 0 | T H E S C IE N T IST 27

Looking forward, looking back Researchers’ understanding of South America’s earliest inhabitants would benefit from the discovery of more human remains, researchers say, but such finds have been hard to come by—which itself raises questions, Dillehay says, of what

GENETIC INSIGHTS ABOUT THE FIRST SOUTH AMERICANS Two studies published in late 2018, one led by David Reich of Harvard University (results depicted in cool colors) and the other by Eske Willerslev of the University of Copenhagen (results in warm colors), compared ancient and modern genomes from across the Americas to infer that there were multiple waves of migration from the northern continent to the southern one.

A wave of migration made up of people genetically related to the famous Clovis culture of North America enters the continent sometime after 17,500 BP. A wave of migrants begins to replace the first wave at around 9,000 BP. A lineage with genetic links to ancient people on the California Channel Islands begins to expand in the Andes by 4,200 BP. A first wave of migration occurred around 14,000 BP. Later, people who had diverged from North American groups and spent time in Central America migrated south. 28 T H E SC I EN TIST | the-scientist.com

happened to the bodies of ancient people. “One major thing is to understand the mortuary practices [of ancient people in the Americas] a little better,” he says. With or without remains, more archaeological sites in parts of Central America and the northwest coast of South America are needed to reveal how people migrated onto and within the continent, and how many waves of migration there were, Boëda notes. The oldest widely accepted archaeological sites in Central America date to around 11,500 BP, and the oldest

ILLUSTRATION BY © JULIA GALOTTA; DATA TAKEN FROM CELL, 175:1185–97.E22, 2018; SCIENCE, 362:EAAV2621, 2018.

mounds] came from . . . and which language they used to speak,” says Hünemeier, who is now working with ancient DNA to try to learn more about the prehistoric coastal group.

Thanks to newer technologies and approaches, archaeologists can now mine more information from sites once they’re identified.

human remains in the area to about 6,000 BP. The region’s acidic soil inhibits preservation of human remains, and some archaeologists suspect that rising sea levels at the end of the Last Glacial Maximum may have swallowed some of the earliest signs of human activity. Thanks to newer technologies and approaches, archaeologists can now extract more information from sites once they’re identified. These methods include genome analysis and stable isotope analysis, which allows researchers to determine whether an individual’s early diet was made up of plants and animals from the local area or from further afield. This in turn can reveal whether a person was from the area where their remains were found or had migrated there. With these new methods, says Rademaker, “each [new] site is going to tell us something incredibly useful.” g

References 1. D.H. Sandweiss et al., “Quebrada Jaguay: Early South American maritime adaptations,” Science, 281:1830–32, 1998. 2. K. Rademaker et al., “Paleoindian settlement of the high-altitude Peruvian Andes,” Science, 346:466–69, 2014. 3. T.D. Dillehay, M.B. Collins, “Early cultural evidence from Monte Verde in Chile,” Nature, 332:150–52, 1988. 4. T.D. Dillehay et al., “New archaeological evidence for an early human presence at Monte Verde, Chile,” PLOS ONE, 10:e0141923, 2015. 5. E. Boëda et al., “New data on a Pleistocene archaeological sequence in South America: Toca do Sítio do Meio, Piauí, Brazil,” PaleoAmerica, 2:286–302, 2016. 6. R.A. Fariña et al., “Arroyo del Vizcaíno, Uruguay: a fossil-rich 30-ka-old megafaunal locality with cut-marked bones,” Proc R Soc B, 281:20132211, 2014. 7. A. Strauss et al., “Early Holocene ritual complexity in South America: the archaeological record of Lapa do Santo (east-central Brazil),” Antiquity, 90:1454–73, 2016. 8. C. Posth et al., “Reconstructing the deep population history of Central and South America,” Cell, 175:1185–97.e22, 2018. 9. J.V. Moreno-Mayar et al., “Early human dispersals within the Americas,” Science, 362:eaav2621, 2018. 10. J. Lindo et al., “The genetic prehistory of the Andean highlands 7000 years BP though European contact,” Sci Adv, 4:eaau4921, 2018. 11. N. Nakatsuka et al., “A paleogenomic reconstruction of the deep population history of the Andes,” Cell, 181:1131–45.e21, 2020. 12. M.A. Castro e Silva et al., “Genomic insight into the origins and dispersal of the Brazilian coastal natives,” PNAS, 117:2372–77, 2020.

CREDIT LINE

WHERE EARLY SOUTH AMERICAN MIGRANTS CAME FROM The widely, though not universally, held thinking among researchers in the field is that indigenous Americans came from East Asia at a time when sea levels would have been low enough to form an inhabitable region known as Beringia that spanned what’s now eastern Siberia and western Alaska. According to the Beringian standstill hypothesis, glaciers long prevented movement further into the Americas, so the founding population remained in the region for thousands of years. When the ice melted enough to permit passage, the Beringians moved fast. “I think we have pretty compelling evidence from the Y DNA studies and the mtDNA [mitochondrial DNA] studies that roughly between 15,400 or so years ago and maybe 14,300 . . . we have an expansion of lineages,” says Ben Potter, an archaeologist at Liaocheng University in China. “We have a star-like radiation” of groups that moved into different areas of North America. A caveat, he says, is that no human remains from this period have been found, and there is thus no fossil evidence to show where exactly the earlier initial expansion originated or where the genetic isolation took place. Some studies have challenged the majority view that that expansion represented humans’ first foray into the Americas. In a 2017 Nature paper, for example, paleontologist Kathleen Holen, then at the San Diego Natural History Museum, and colleagues reported finding 130,000-year-old mastodon bones at a site in California that appeared to have been smashed with human-made stone tools (544:479–83). The study was met with skepticism by researchers who said the bones could instead have been broken by other means, such as modern construction equipment. In July of this year, an international team led by Eske Willerslev of the University of Copenhagen garnered headlines with another report in Nature, this one of purportedly human-made tools dating to 33,000 BP in a cave in Mexico (doi:10.1038/s41586-020-2509-0). But critics questioned whether the stones in question had truly been shaped by humans.  Another disputed idea, known as the Solutrean hypothesis, is based on similarities between tools made by North America’s early Clovis culture and those made by Europe’s Solutrean people between 22,000 and 17,000 BP. Its main proponents, Bruce Bradley of the University of Exeter and the late Dennis Stanford of the Smithsonian National Museum of Natural History, suggested that some Solutreans could have migrated to North America via boats that hugged an ice bridge between the two continents (World Archaeol, 36:459–78, 2004). Other researchers have pointed out what they see as multiple problems with the hypothesis, including the 7,000-year gap between Solutreans’ use of the tools in Europe and the first Clovis site in North America, and a lack of genetic evidence that any early Americans had European ancestry. The Beringian standstill hypothesis continues to undergo revisions as new studies emerge. In 2018, for example, Potter and his colleagues reported a genetic analysis of 11,500-year-old remains of a baby girl found buried in Alaska at a site called Upward Sun. The infant belonged to a previously unknown lineage that split from the ancestors of today’s Native Americans between 22,000 and 18,100 years ago, the researchers suggested (Nature, 553:203–207). Potter notes that the potential for interbreeding between known populations and extinct lineages such as this can complicate genetic models that estimate how long ago populations diverged, and he says he thinks future discoveries like that made at Upward Sun are likely. “There probably are more of those out there that we just haven’t detected yet,” he says. 09. 2020 | T H E S C IE N T IST 2 9

Title Here in Serif

Becoming Sapiens

BY AUTHOR HERE

S

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30 T H E SC I EN TIST | the-scientist.com

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© MARK GARLICK, SCIENCE PHOTO LIBRARY

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t’s not unusual for geochronologist Rainer Grün to bring human bones back with him when he returns home to Australia from excursions in Europe or Asia. Jawbones from extinct hominins in Indonesia, Neanderthal teeth from Israel, and ancient human finger bones unearthed in Saudi Arabia have all at one point spent time in his lab at Australian National University before being returned home. Grün specializes in developing methods to discern the age of such specimens. In 2016, he carried with him a particularly precious piece of cargo: a tiny sliver of fossilized bone covered in bubble wrap inside a box. The bone fragment had come from a skull—still stored at the Natural History Museum in London—with a heavy brow ridge and a large face. It looked so primitive that the miner who had discovered it in 1921 at a lead mine in the Zambian town of Kabwe, then in the British territory of Rhodesia, first thought it had belonged to a gorilla. But later that year, museum paleontologist Arthur Smith Woodward noticed what he interpreted as typically human features, such as the skull’s thin and relatively large braincase, that motivated him to designate the specimen as its own hominin species. In the 1980s, however, museum paleoanthropologist Chris Stringer took another look at the skull and classified it as belonging to the species Homo heidelbergensis, an ancient hominin thought to be a human ancestor. Based on its primitiveness, Stringer says, most researchers guessed it was an early individual who lived around half a million years ago, some 200,000 years before the earliest Homo sapiens were starting to emerge. But nobody knew exactly how old the skull was. For decades, no dating method existed that could identify the fossil’s age without the destructive process of grinding up bits of bone for analysis. But Grün was determined to find a solution. Grün is one of very few geochronologists proficient in a laser technique that extracts and reduces a barely visible grain of bone— smaller than the bone’s natural pores—to atoms, he says. The laser is coupled with a

mass spectrometer, which measures the concentrations of uranium isotopes that undergo radioactive decay at a specific rate over time. Having returned from his trip to procure the Homo heidelbergensis sample, Grün watched as the laser poked two tiny holes into the bone fragment and the particles disappeared into the mass spectrometer. Upon evaluating the mass spec data, he could tell that the fragment was much younger than previously believed. As he, Stringer, and others reported in Nature this past April, their best estimate was 299,000 years, give or take 25,000.1 That meant that the Kabwe individual had lived not before, but around the same time as the first Homo sapiens–like people dwelled in North Africa. Along with other archaeological evidence, the findings suggest that perhaps Homo heidelbergensis was not our ancestor, but a neighbor. Together with yet another hominin, Homo naledi, known to have existed in southern Africa at that time, Africa may have been a crowded place. “Ten years ago, I think most of us would’ve thought, well, Africa in the last 300,000 years is just going to show you the evolution of Homo sapiens, and that’s really all—the other species would have disappeared, gone extinct,” notes Stringer. “Now we know that there were probably at least three different kinds of hominins around.” That’s akin to the situation that unfolded in Eurasia, where Neanderthals and Denisovans thrived for hundreds of thousands of years before Homo sapiens migrated out of Africa and at times even interbred with the other hominin groups. The story in Africa remains murky, however, as researchers have not been able to reconstruct human history in vivid detail, in part because hominin fossils informative about our species’ emergence and coexistence with other species are rare in Africa. As a result, finds such as the Kabwe skull continue to raise more questions than answers. If Homo heidelbergensis wasn’t one of our recent ancestors, then who was? If our species really did overlap in time with Homo heidelbergensis, what role did they play in our evolutionary history? In recent years, a field that has traditionally relied on fossil discoveries has acquired

helpful new tools: genomics and ancient DNA techniques. Armed with this combination of approaches, researchers have begun to excavate our species’ early evolution, hinting at a far more complex past than was previously appreciated—one rich in diversity, migration, and possibly even interbreeding with other hominin species in Africa. “To piece together that story, we need information from multiple different fields of study,” remarks Eleanor Scerri, an archaeologist at the Max Planck Institute for the Science of Human History in Jena, Germany. “No single one is really going to have all the answers—not genetics, not archaeology, not the fossils, because all of these areas have challenges and limitations.”

A sparse fossil record Bones easily disintegrate in many parts of Africa, in acidic forest soils or dry, sunexposed areas. Moreover, the continent is largely unexplored by archaeologists. While northwestern Africa and former British territories in eastern and southern Africa have a long tradition of professional archaeological research, few researchers have looked for fossils anywhere else, notes archaeologist Khady Niang of Cheikh Anta Diop University in Senegal. That’s especially the case for the western and central parts of the continent, where preservation conditions are also poor and excavations difficult at times due to political instability. “We might be missing some really, really important parts of the story,” adds Yale University anthropologist Jessica Thompson. What African hominin fossils do make clear is the depth of humanity’s roots on that continent. Researchers have found some of the most abundant fossils in sediments between 3.5 million and 3.2 million years old. That appeared to be the heyday of the australopiths (including the genus Australopithecus), apes that walked upright and are believed to have used stone tools, but still climbed trees and had relatively small brains. It’s thought that somehow our own genus, Homo, emerged from transitional ape species some 2.8 million years ago as a clan of hominins with distinctive teeth, probably adapted to an eclectic diet that allowed them to thrive 09. 2020 | T H E S C IE N T IST 3 1

in a wide range of habitats. But there are few sediments, let alone fossils, left behind from that time, making the birth of our genus one of the most poorly understood periods in our evolution, Thompson notes. The fossil record yields more secrets about the time shortly after the emergence of Homo, revealing a diversity of different Homo species in Africa, of which Homo erectus seems to persist the longest. H. erectus crops up in Africa’s limited fossil record around 2 million years ago and hangs around on the continent until roughly a million years ago. It was the first hominin that shows evidence of having lived in human-like social groupings and used fire, and it is thought to be a human ancestor. When and how H. sapiens emerged isn’t at all clear, but what is apparent is that we weren’t alone; fossils suggest that several other hominin species, such as that represented by the Kabwe skull, inhabited the continent at the time our species appeared. Another relatively small-brained hominin, Homo naledi, is also thought to have lived in southern Africa around 300,000 years ago.2,3 And inside a Moroccan cave called Jebel Irhoud, 300,000-year-old skeletons were found that carry very early features of H. sapiens.4 It’s not yet known how long those different hominin species existed, however, or whether they physically overlapped and perhaps even shared genes with one another, Stringer notes, or whether there were others. By around 160,000 years ago, the constellation of physical features that defines us today—such as a globular braincase and a pointed chin—had begun to emerge in ancient hominin groups represented by fossils found across Africa. Later, some of these anatomically modern humans crossed the thin spit of land that connects Africa to Eurasia, probably on several occasions. On that new continent, they eventually met Neanderthals and Denisovans, which, like two hobbit-size Homo species found on southeast Asian islands, are thought to be the evolutionary products of earlier hominin migrations out of the continent. “Africa was this sort of leaky faucet, and hominins were just dribbling out of it all the time,” Thompson says. 32 T H E SCI ENT I ST | the-scientist.com

Fossil finds over the years have steadily bolstered a long-held idea that anatomically modern humans first emerged in Africa. This “Out of Africa” model, proposed by anthropologists in the late 20th century, posited that all humans of Eurasian ancestry descended from a single ancestral African population, which then spread throughout the world and displaced all other hominins. The opposing “multi-regionalism” model, by contrast, conceived that multiple human subpopulations—which stemmed from regional lineages of an ancestral species such as Homo erectus—existed across Europe, Asia, and Africa, and through continuous mixing evolved together to form the present human population. While fossils supported the former theory, it was the advent of genetic research that showed unequivocally that populations outside of Africa descended from a single population in Africa. But the story had a twist: in two groundbreaking studies published in 2014, researchers compared ancient DNA extracted from Neanderthal bones and compared it with modern-day people, and found that 2 percent of the average European genome is Neanderthal in origin.5,6 Our species originated in Africa, but interbred with hominins outside of it. (See “Our Inner Neanderthal,” The Scientist, September 2019.) These findings, and many since, have highlighted the power of genetics in resolving questions about human ancestry that fossils alone cannot. Investigations of the genomes of living Africans are now underway to help fill in the gaps of Africa’s fossil record. “[Such studies] are really providing important insights into our population history and African origins,” says Yale University evolutionary biologist Serena Tucci. “We are getting to know and understand processes that happened very early on in our evolutionary history.”

Ghost hominins Even the very first investigations of our genetic ancestry, gleaned from small, bitesize chunks of genetic material, positioned Africa as the cradle of humanity. One widely publicized 1987 study compared mitochondrial gene snippets from 147

people across the world, and concluded that Africans have the highest mitochondrial diversity, suggesting that our species originated and spent most of its evolutionary history there.7 Specifically, the authors traced all human mitochondrial diversity back to a single theoretical woman who lived in East Africa hundreds of thousands of years ago, whom the media popularized as “mitochondrial Eve.” Later studies estimated that the most recent common ancestor of modern Y chromosome variation (dubbed “Y chromosome Adam”) could also be traced back to Africa. Subsequent studies of nuclear DNA have validated our African birthplace and refined our knowledge of the human genetic landscape. Several studies of genetic variation among modern-day Khoe and San individuals, two groups of indigenous people in southern Africa known for their click language, have suggested they represent our species’ most genetically diverse lineage.8 Collectively known as Khoe-San, this group is thought to have split from other populations between 200,000 and 350,000 years ago, making them the most ancient population of modern humans to diverge. NonAfricans, meanwhile, represent a reduced subset of the diversity in Africa, and likely trace most of their ancestry back to just one small population—probably no more than a few thousand individuals—who ventured out of the continent between 60,000 and 70,000 years ago.9 Some scientists see the extraordinary diversity in modern Khoe-San people as evidence that our species arose in southern Africa. Along with some archaeological evidence from the region, that challenges the long-held idea of an East African origin, which was based on the fact that many early hominin fossils were found there. However, trying to pinpoint the precise location of our species’ origins from DNA is often criticized for the simple reason that people move around— it’s not known if the populations living in one place today were there hundreds or thousands of millennia ago. In fact, some researchers, including Scerri, Stringer, and Thompson, have recently constructed

an entirely new theory of our origins: that anatomically modern humans didn’t arise from a single place, but gradually emerged from a web of interconnected populations sprawled across Africa—a continental gene-sharing bonanza that hominin lineages besides our own may have participated in.10 “It’s a good way to interpret the data we have right now,” says Niang. In addition to where we evolved, researchers are interested in how: which genes gave us a selective advantage to survive in particular environments, and which ancestors contributed to our genomes? Unfortunately, modern African DNA is severely underrepresented in genetic research, making these questions particularly challenging to answer. Most sequenced genomes are of European origin, with fewer than 2 percent coming from Africans. This dearth of African genomes is compounded by the fact that the genetic scaffold underlying some frequently studied traits such as skin pigmentation appear to be far more complex in Africans than in other populations, notes Brenna Henn, a population geneticist at the University of California, Davis. “The twelve to fifteen genes [for skin pigmentation] that people cite in Eurasian populations explain less than 25 percent of the variation in Africans.”

tricky to distinguish that older history when there’s been this newer wave of gene flow messing with your modeling.” Still, geneticists have been able to tease out some signals from our distant past, using computational models that ask what kind of evolutionary processes—such as mutation, selection, and interbreeding with other groups—best explain the pattern of variation across modern genomes. One intriguing finding of such studies is possible evidence of mixture with now-extinct, unknown groups of modern humans and other hominins: “ghost” populations that, like Neanderthals, left traces in modern genomes. In one analysis of 15 sequenced genomes, Tishkoff ’s group investigated the sources of genetic variation in three different modern African hunter-gatherer groups.11 The team’s models suggested that interbreeding with an archaic hominin species—which seemed as different from modern humans as are Neanderthals—was the most likely origin for a set of unusual sequences they found. “The model that includes a ghost population is always better [to fit the data], basically,” Tishkoff says. A handful of similar studies have also revealed traces of ghost hominins in modern African genomes, sometimes account-

With few sediments, let alone fossils, left behind from that time, the birth of our genus is one of the most poorly understood periods in our evolution. African population history complicates matters further. Large-scale migrations pulled people back and forth across the continent for thousands of years. People from Eurasia also migrated back to Africa. Where people moved, they swapped their genes with local populations, shuffling patterns of ancestry across African genomes. This upheaval of ancient population structures creates one of the biggest challenges in teasing out archaic history from modern genomes, notes University of Pennsylvania geneticist Sarah Tishkoff. “It can make it very

ing for up to 10 to 20 percent of the genetic variation. Some research suggests that mixing took place after the ancestors of modern Eurasians left Africa, hinting that other kinds of hominins could have existed alongside Homo sapiens in Africa until very recently. “It’s actually pretty convincing,” says Henn, who wasn’t involved in these studies. “Ten percent of the genome—I’m going to have a hard time invoking one single other process that can explain a signal like that.” Ultimately, researchers need samples of DNA from ancient hominins to prove whether archaic African species did in fact contribute

to modern genetic variation. While scientists have managed to overcome some of the technical hurdles of sequencing highly degraded ancient DNA from human fossils in Africa, the oldest human DNA found on the continent is just 15,000 years old, an age that pales in comparison to some 400,000-year-old hominin DNA found in a cave in Spain with relatively cool, stable temperatures. Archaeologists can only dream of finding intact DNA that old on the African continent, notes Tessa Campbell, an ancient DNA specialist at Iziko Museums of South Africa. “No one wants to say never . . . but it’s very unlikely.” Because DNA is unlikely to survive very long in the African heat, researchers have largely refrained from drilling into the fossils they’ve found of other hominins in Africa for fear of destroying them. But efforts are underway to study ancient DNA from younger fossils of Homo sapiens to crack other mysteries about human history on the continent, Tucci notes. “This is definitely a new era for African genomics.”

Mining bones for ancient DNA In 2015, an international team of researchers managed to harvest the first ancient DNA in Africa—the genome of Mota, a man who left behind 4,500-year-old remains in an Ethiopian cave.12 In the five years since that publication, researchers have published nearly 100 other full and partial ancient human sequences from Africa. These genomes have helped scientists better understand the messy signatures from recent migration events that make studies of modern genomes so difficult. For instance, mitochondrial DNA from the skulls of seven people who lived some 15,000 years ago in modern-day Morocco revealed that they were closely related to Natufians, hunter-gatherers who dwelled in the Near East, as well as people living south of the Sahara desert.13 This finding suggested that there were far-flung connections between North Africa, the Near East, and sub-Saharan Africa before the dawn of agriculture. Analyses of ancient DNA have also helped researchers understand how ancient migrations affected the genomes of people alive today. One such migration is the Bantu 09. 202 0 | T H E S C IE N T IST 3 3

lier back-migrations into the continent and eventually carried to the southernmost tip of Africa as other migrating human populations moved southward, the researchers found. Such studies have also provided insight into deep divergences that occurred in human populations long before migrations of farmers and herders. Mary Prendergast, an anthropologist at Saint Louis University in Madrid, and her colleagues recently sequenced the first ancient DNA from West Africa, material extracted from the remains

of children buried inside a rock shelter in Cameroon.15 Comparing the 3,000- and 8,000-year-old DNA with ancient genomes collected elsewhere and with genomes of modern people allowed the researchers to reconstruct some of the earliest branches of our species’ evolutionary tree. In addition to the deep split between Khoe-San groups and other African populations— from which non-Africans also descend— their model suggested that two other major lineages split just as deeply, diverging from

OUR HISTORY IN AFRICA

Hominin fossils that reveal clues to the emergence of Homo sapiens are rare in Africa, but in combination with studies of modern human genomes, researchers are piecing together an ever more complex timeline of human history.

HOMO HEIDELBERGENSIS Named after its initial discovery near Heidelberg, Germany, fossils similar to Homo heidelbergensis were later found to also occur in Africa. It routinely hunted large animals and may have built dwellings made of wood or rock. HOMO ERECTUS Homo erectus may have been the first hominin to wield fire and stone axes. The species spread across Asia, where it continued to evolve. In Africa, it gave rise to Homo heidelbergensis.

*Note that the scale of this graphic changes across its width. While Homo sapiens has existed for the past few hundred thousand years, it’s noteworthy that Homo erectus, living from around 2 mya to perhaps 100 kya, is still the longest-lived hominin species.

EARLY HOMO SPECIES Hominins of the genus Homo shared distinctly small back teeth, which experts think allowed them to consume diverse diets. One species of early Homo evolved into Homo erectus.

SAHELANTHROPUS, ORRORIN, ARDIPITHECUS SPP. Members of these relatively small-brained genera probably emerged not long after the human-chimpanzee divergence, and are the first known species of apes that habitually walked upright.

7 mya

6

AUSTRALOPITHECUS SPP. This diverse group exhibited both ape- and human-like characteristics. Some species are thought to have used stone tools and to have evolved into the genus Homo.

5

4

3

2

1 mya

© ISTOCK.COM, GIORGIOMORARA

expansion, which gradually spread West African farming practices across the continent between roughly 5,000 and 1,000 years ago. By comparing DNA from ancient huntergatherer remains in southern Africa with modern-day Khoe-San people, evolutionary biologist Carina Schlebusch of Uppsala University in Sweden and her colleagues found that some Khoe-San groups carry DNA that ancient farmers brought with them.14 They also carry mixed Eurasian ancestry that had been introduced to North Africa with ear-

It was not a streamlined process of australopiths steadily evolving into modern humans, but a messy and haphazard journey that includes interwoven ancestries of many groups.

one another more than 200,000 years ago. One lineage is ancestral to central African hunter-gatherers known as Aka and Mbuti, and the second is a previously unknown

“ghost” lineage whose fate is uncertain. “There’s all this deep, deep population structure with various differentiated branches of the human tree throughout the Pleistocene

in Africa that we haven’t really appreciated very much yet,” Prendergast says. Only time will tell whether researchers’ current arsenal of technologies is enough to untangle the complete story of human evolution. Perhaps novel technologies—such as paleoproteomics, a nascent field that aims to reconstruct ancestry from fossilized proteins, which are more durable than DNA—will help researchers “push further back in time,” notes biological anthropologist Rebecca Ackermann of the University of Cape Town.

Comparing ancient DNA from Neanderthals and Denisovans with modern human genomes has revealed that modern humans interbred with these other hominin groups.

DENISOVANS Denisovans, only known from from ancient DNA and a handful of bones and teeth, were closely related to Neanderthals.

Genomic analyses suggest that the majority of people living outside Africa today trace most of their ancestry back to a single migration event of a small group of modern humans who left Africa between 60,000 and 70,000 years ago.

HOMO NEANDERTHALENSIS Compared with modern humans, Neanderthals had shorter, stockier skeletons and larger noses, but their brains were just as large, if not larger.

HOMO SAPIENS Eventually, the hallmarks of our own species—exceptionally large brains, flat faces, and small jaws— appear in the fossil record. Genetic studies suggest that H. sapiens started to split into several major lineages of modern humans more than 200,000 years ago.

HOMO NALEDI Only known from skeletons found in South Africa, this species had a remarkably tiny brain but modern-human–like features such as the shape of its teeth and possibly the habit of burying its dead.

Some analyses of modern human genomes hint that Homo sapiens may have interbred with other hominins in Africa.

500 kya

100 kya

HOMO FLORESIENSIS AND HOMO LUZONENSIS Researchers have recently discovered two small hominin species on Pacific islands, Homo floresiensis on Flores in Indonesia and Homo luzonensis in the Philippines.

50 kya

Present

EXCAVATING A CONTINENT

AFAR REGION, ETHIOPIA, 2013

Lucy—the skeletal remains of an Australopithecus afarensis female—is one of the best-known hominin fossils. Studies suggest that she was both tree-dwelling and capable of an upright gait, providing an important evolutionary stepping stone from more primitive ape species to modern humans.

Adult jawbone, 2.8 million years ago

NEAR SAFI, MOROCCO, 1961

Human remains at Jebel Irhoud, 315,000 years ago

A mandible fragment is the earliest known trace of the genus Homo, although the species it belongs to is a mystery.

Flint blades and Homo sapiens–like skeletons in a Moroccan cave known as Jebel Irhoud may represent the oldest Homo sapiens artifacts. The skeletons have modern features such as round skulls and modern-human–like teeth and faces.

OMO NATIONAL PARK, ETHIOPIA, 1967-1974

Omo Kibish remains, 195,000 years ago

Fragments from two skulls, four jaws, a legbone, a few hundred teeth, and some other bones were found at a site in Ethiopia, and are classified as anatomically modern Homo sapiens.

LAKE TURKANA, KENYA, 1984

“Turkana Boy,” 2 million years ago A nearly complete skeleton of an ancient Homo erectus child found near Kenya’s Lake Turkana provides a rare glimpse into how quickly this species reached adulthood and how similar their skeletons were to ours.

RISING STAR CAVE, SOUTH AFRICA, 2013

Homo naledi, 236,000–335,000 years ago

KABWE, ZAMBIA, 1921

“Kabwe skull,” 300,000 years ago Also called “Broken Hill skull,” the specimen is considered a representative of Homo heidelbergensis.

36 T H E SC I EN TIST | the-scientist.com

In 2013 and 2014, cavers found skeletons of two adults and one juvenile of what is believed to be a new species: Homo naledi. Its tiny brain and ape-like shoulders—indicating it was a good climber—suggest it may be an evolutionary offshoot lineage that went extinct.

WIKIMEDIA, 120; ARIZONA STATE UNIVERSITY; WIKIMEDIA, BAHN, PAUL G; WIKIMEDIA, GERBIL; WIKIMEDIA, RYAN SOMMA; ELIFE, 6:E24232, 2017; WIKIMEDIA, GUILLAUMEG

A number of researchers suspect that Homo sapiens arose not in a single place in Africa, but across the entire continent, emerging from a network of interconnected hominin populations. But for decades, archaeologists positioned East and South Africa as important places for hominin evolution and the putative birthplace of our species. That’s likely because most fossils, including groundbreaking findings that have transformed our understanding of human evolution, have been AFAR REGION, ETHIOPIA, 1974 found in those regions. “Lucy,” 3.2 million years ago

What is already abundantly clear is that human evolution was far more complex than previously appreciated by anthropologists. It was not a streamlined process of australopiths steadily evolving into modern humans, but a messy and haphazard journey that includes interwoven ancestries of many groups, some of which have never been discovered other than through the genetic traces they left in ancient and modern genomes. “We have a long history. A lot of things happened, and a lot of ancestors contributed to our genomes today,” Schlebusch says. “It’s not going to be a simple story.” g Katarina Zimmer is a New York–based freelance journalist. Find her on Twitter @katarinazimmer.

References 1. R. Grün et al., “Dating the skull from Broken Hill, Zambia, and its position in human evolution,” Nature, 580:372–75, 2020. 2. L.R. Berger et al., “Homo naledi, a new species of the genus Homo from the Dinaledi Chamber, South Africa,” eLife, 4:e09560, 2015. 3. P.H. Dirks et al., “The age of Homo naledi and associated sediments in the Rising Star Cave, South Africa,” eLife, 6:e24231, 2017. 4. J.-J. Hublin et al., “New fossils from Jebel Irhoud, Morocco and the pan-African origin of Homo sapiens,” Nature, 546:289–92, 2017. 5. B. Vernot, J.M. Akey, “Resurrecting surviving Neanderthal lineages from modern human genomes,” Science, 343:1017–21, 2014. 6. S. Sankaraman et al., “The genomic landscape of Neanderthal ancestry in present-day humans,” Nature, 507:354–57, 2014. 7. R.L. Cann et al., “Mitochondrial DNA and human evolution,” Nature, 325:31–36, 1987. 8. C.M. Schlebusch et al., “Genomic variation in seven Khoe-San groups reveals adaptation

and complex African history,” Science, 338:374–79, 2012. 9. S. Tucci, J.M. Akey, “A map of human wanderlust,” Nature, 538:179–80, 2016. 10. E.M.L. Scerri et al., “Did our species evolve in subdivided populations across Africa, and why does it matter?” Trends Ecol Evol, 33:582–94, 2018. 11. J. Lachance et al. “Evolutionary history and adaptation from high-coverage wholegenome sequences of diverse African huntergatherers,” Cell, 150:457–69, 2012. 12. M.G. Llorente et al., “Ancient Ethiopian genome reveals extensive Eurasian admixture in Eastern Africa,” Science, 350:820–22, 2015. 13. M. v.d. Loosdrecht et al., “Pleistocene North African genomes link Near Eastern and sub-Saharan African human populations,” Science, 360:548–52, 2018. 14. C.M. Schlebusch et al., “Southern African ancient genomes estimate modern human divergence to 350,000 to 260,000 years ago,” Science, 358:652–55, 2017. 15. M. Lipson et al., “Ancient West African foragers in the context of African population history,” Nature, 577:665–70, 2020.

CREDIT LINE

DECOLONIZING STUDIES OF HUMAN EVOLUTION The San people of southern Africa are one of the most intensively studied indigenous groups in the world. Their click language and traditional hunter-gatherer lifestyles have long fascinated anthropologists. And the antiquity of their genetic lineage makes them a treasure trove for geneticists studying human evolutionary history. However, studies on San lifestyles and genomes have not always been conducted ethically. For instance, scientists have sometimes referred to the San as “bushmen,” a derogatory term associated with colonial-era researchers using modern indigenous groups as models of primitive human ancestors, and have taken photographs of children and breastfeeding mothers without permission. “We’re not saying that everybody is bad. But you get those few individuals who don’t respect the San,” Leana Snyders, head of the South African San Council in Upington, South Africa, told Science in 2017. Ethical conduct in genomic research came to the foreground in 2010 following a high-profile analysis of San genomes in Nature in which the authors had, among other transgressions, not asked San leaders for permission to conduct the study.   All disciplines that study human evolution in Africa have at times been criticized for their extractive nature. Archaeological research—a field pioneered by European colonial nations—has long been driven by Western researchers digging up fossils from Africa to study them, sometimes taking them elsewhere to do so. Some hominin fossils are still displaced, such as the Kabwe skull, a famous Homo heidelbergensis specimen that remains in London’s Museum of Natural History, despite Zambia’s multiple requests to repatriate the skull. According to an April press release, the museum has approached Zambian authorities to begin discussing the possible return of the skull following a 2018 agreement between the UK and Zambia to find a solution to the issue. Some scientists have called for regulations to protect fossil collections from ancient DNA research, whereby African hominin fossils undergo the damaging process of extracting DNA. Now, “African museums are taking a leading role to make sure this [research] happens through collaboration and regulation,” notes anthropologist Mary Prendergast of Saint Louis University’s Madrid campus, as geneticists are working to develop new, less destructive techniques for ancient DNA analysis. The San, for their part, created a code of research conduct in 2017 that, for example, requires researchers to respect their communities and to allow them to comment on findings prior to publication to avoid derogatory interpretations. Researchers are also required to compensate the community for their cooperation, through financial support, knowledge, or job opportunities, for instance. A number of scientists have called for a greater role of African scientists in human evolutionary research. To make that possible, Western funding agencies and institutions have an obligation to support African efforts to improve their countries’ antiquities infrastructure, so that “the next generation of African scholars [can] take control of the research in their areas,” notes anthropologist Eleanor Scerri of the Max Planck Institute in Germany. Foreign research teams should also foster stronger collaboration with African researchers, rather than simply seeking their help with fossil excavations, which has sometimes been the case, notes University of Cape Town biological anthropologist Rebecca Ackermann. Research groups have become more diverse, she notes, but the transition is slow. “I do see a change. It’s just not as fast as I would like.” 09. 2020 | T H E S C IE N T IST 37

The Making of an Organism Understanding biology’s software—the rules that enable great plasticity in how cell collectives build organs and organisms—is key to advancing tissue engineering and regenerative medicine.

© ISTOCK.COM, LUCKYSTEP48

BY MICHAEL LEVIN

40 T H E SC I EN TIST | the-scientist.com

multicellularity into a new form, without genomic changes.1 This represents an extremely exciting sandbox in which bioengineers can play, with the aim of decoding the logic of anatomical and behavioral control, as well as understanding the plasticity of cells and the relationship of genomes to anatomies. Deciphering how an organism puts itself together is truly an interdisciplinary undertaking. Resolving the whole picture will involve understanding not only the mechanisms by which cells operate, but also elucidating the computations that cells and groups of cells carry out to orchestrate tissue and organ construction on a whole-body scale. The next generation of advances in this area of research will emerge from the flow of ideas between computer scientists and biologists. Unlocking the full potential of regenerative medicine will require biology to take the journey computer science has already taken, from focusing on the hardware—the proteins and biochemical pathways that carry out cellular operations— to the physiological software that enables networks of cells to acquire, store, and act on information about organ and indeed whole-body geometry. In the computer world, this transition from rewiring hardware to reprogramming the information flow by changing the inputs gave rise to the information technology revolution. This shift of perspective could transform biology, allowing scientists to achieve the still-futuristic visions of regenerative medicine. An understanding of how independent, competent agents such as cells cooperate and compete toward robust outcomes, despite noise and changing environmental conditions, would also inform engineering. Swarm robotics, Internet of Things, and even the development of general artificial intelligence will all be enriched by the ability to read out and set the anatomical states toward which cell collectives build, because they share a fundamental underlying problem: how to control the emergent outcomes of systems composed of many interacting units or individuals.

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E

fforts to use regenerative medicine—which seeks to address ailments as diverse as birth defects, traumatic injury, aging, degenerative disease, and the disorganized growth of cancer— would be greatly aided by solving one fundamental puzzle: How do cellular collectives orchestrate the building of complex, three-dimensional structures? While genomes predictably encode the proteins present in cells, a simple molecular parts list does not tell us enough about the anatomical layout or regenerative potential of the body that the cells will work to construct. Genomes are not a blueprint for anatomy, and genome editing is fundamentally limited by the fact that it’s very hard to infer which genes to tweak, and how, to achieve desired complex anatomical outcomes. Similarly, stem cells generate the building blocks of organs, but the ability to organize specific cell types into a working human hand or eye has been and will be beyond the grasp of direct manipulation for a very long time. But researchers working in the fields of synthetic morphology and regenerative biophysics are beginning to understand the rules governing the plasticity of organ growth and repair. Rather than micromanaging tasks that are too complex to implement directly at the cellular or molecular level, what if we solved the mystery of how groups of cells cooperate to construct specific multicellular bodies during embryogenesis and regeneration? Perhaps then we could figure out how to motivate cell collectives to build whatever anatomical features we want. New approaches now allow us to target the processes that implement anatomical decision-making without genetic engineering. In January, using such tools, crafted in my lab at Tufts University’s Allen Discovery Center and by computer scientists in Josh Bongard’s lab at the University of Vermont, we were able to create novel living machines, artificial bodies with morphologies and behaviors completely different from the default anatomy of the frog species (Xenopus laevis) whose cells we used. These cells rebooted their

(Re)Building a body Many types of embryos can regenerate entirely if cut in half, and some species are proficient regenerators as adults. Axolotls (Ambystoma mexicanum) regenerate their limbs, eyes, spinal cords, jaws, and portions of the brain throughout life. Planarian flatworms (class Turbellaria), meanwhile, can regrow absolutely any part of their body; when the animal is cut into pieces, each piece knows exactly what’s missing and regenerates to be a perfect, tiny worm. The remarkable thing is not simply that growth begins after wounding and that various cell types are generated, but that these bodies will grow and remodel until a correct anatomy is complete, and then they stop. How does the system identify the correct target morphology, orchestrate individual cell behaviors to get there, and determine when the job is done? How does it communicate this information to control underlying cell activities? Several years ago, my lab found that Xenopus tadpoles with their facial organs experimentally mixed up into incorrect positions still have largely normal faces once they’ve matured, as the organs move and remodel through unnatural paths.2 Last year, a colleague at Tufts came to a similar conclusion: the Xenopus genome does not encode a hardwired set of instructions for the movements of different organs during metamorphosis from tadpole to frog, but rather encodes molecular hardware that executes a kind of “error minimization loop,” comparing the current anatomy to the target frog morphology and working to progressively reduce the difference between them.3 Once a rough spatial specification of the layout is achieved, that triggers the cessation of further remodeling. The deep puzzle of how competent agents such as cells work together to pursue goals such as building, remodeling, or repairing a complex organ to a predetermined spec is well illustrated by planaria. Despite having a mechanistic understanding of stem cell specification pathways and axial chemical gradients, scientists really don’t know what determines the intri-

cate shape and structure of the flatworm’s head. It is also unknown how planaria perfectly regenerate the same anatomy, even as their genomes have accrued mutations over eons of somatic inheritance. Because some species of planaria reproduce by fission and regeneration, any mutation that doesn’t kill the neoblast—the adult stem cell that gives rise to cells that regenerate new tissue—is propagated to the next generation. The worm’s incredibly messy genome shows evidence of this process, and cells in an individual planarian can have different numbers of chromosomes. Still, fragmented planaria regenerate their body shape with nearly 100 percent anatomical fidelity. So how do cell groups encode the patterns they build, and how do they know to stop once a target anatomy is achieved? What would happen, for example, if neoblasts from a planarian species with a flat head were transplanted into a worm of a species with a round or triangular head that had the head amputated? Which shape would result from this het-

largely drives clinical approaches, recent literature supports a view of cancer as cells simply not being able to receive the physiological signals that maintain the normally tight controls of anatomical homeostasis. Cut off from these patterning cues, individual cells revert to their ancient unicellular lifestyle and treat the rest of the body as external environment, often to ruinous effect. If we understand the mechanisms that scale single-cell homeostatic setpoints into tissue- and organ-level anatomical goal states and the conditions under which the anatomical error reduction control loop breaks down, we may be able to provide stimuli to gain control of rogue cancer cells without either gene therapy or chemotherapy.

Bioelectrical software: Beyond the brain The software of life, which exploits the laws of physics and computation, is enabled by chemical, mechanical, and electrical signaling across cellular networks. (See illustra-

Deciphering how an organism puts itself together is truly an interdisciplinary undertaking.

erogeneous mixture? To date, none of the high-resolution molecular genetic studies of planaria give any prediction for the results of this experiment, because so far they have all focused on the cellular hardware, not on the logic of the software— implemented by chemical, mechanical, and electrical signaling among cells—that controls large-scale outcomes and enables remodeling to stop when a specific morphology has been achieved. Understanding how cells and tissues make real-time anatomical decisions is central not only to achieving regenerative outcomes too complex for us to manage directly, but also to solving problems such as cancer. While the view of cancer as a genetic disorder still

tion on page 43.) While the chemical and mechanical mechanisms of morphogenesis have long been appreciated by molecular and cell biologists, the role of electrical signaling has largely been overlooked. But the same reprogrammability of neural circuits in the brain that supports learning, memory, and behavioral plasticity applies to all cells, not just neurons. Indeed, bacterial colonies can communicate via ionic currents, with recent research revealing brain-like dynamics in which information is propagated across and stored in a kind of proto-body formed by bacterial biofilms. So it should really come as no surprise that bioelectric signaling is a highly tractable component of morphological outcomes in multicellular organisms. 09. 202 0 | T H E S C IE N T IST 41

A few years ago, we studied the electrical dynamics that normally set the size and borders of the nascent Xenopus brain, and built a computer model of this process to shed light on how a range of various brain defects arise from disruptions to this bioelectric signaling. Our model suggested that specific modifications with mRNA or small molecules could restore the endogenous bioelectric patterns back to their correct layout. By using our computational platform to select drugs to open existing ion channels in nascent neural tissue 4 or even a remote body tissue, 5 we were able to prevent and even reverse brain defects caused not only by chemical teratogens—compounds that disrupt embryonic development—but by mutations in key neurogenesis genes. Similarly, we used optogenetics to stimulate electrical activity in various somatic cell types to trigger regeneration of an entire tadpole tail 6—an appendage with spinal cord, muscle, and peripheral innervation—and to normalize the behavior of cancer cells in tadpoles strongly expressing human oncogenes such as KRAS mutations. 7 We used a similar approach to trigger posterior regions, such as the gut, to build an entire frog eye.8 In both the eye and tail cases, the information on how exactly to build these complex structures, and where all the cells should go, did not have to be specified by the experimenter; rather, they arose from the cells themselves. Such findings reveal how ion channel mutations result in numerous human developmental channelopathies,9 and provide a roadmap for how they may be treated by altering the bioelectric map that tells cells what to build. We also recently found a striking example of such reprogrammable bioelectrical software in control of regeneration in planaria. In 2011, we discovered that an endogenous electric circuit establishes a pattern of depolarization and hyperpolarization in planarian fragments that regulate the orientation of the anterior-posterior axis to be rebuilt.10 Last year, we discovered that this circuit 42 T H E SC I EN TIST | the-scientist.com

controls the gene expression needed to build a head or tail within six hours of amputation, 11 and by using molecules that make cell membranes permeable to certain ions to depolarize or hyperpolarize cells, we induced fragments of such worms to give rise to a symmetrical two-headed form, despite their wildtype genomes. Even more shockingly, the worms continued to generate twoheaded progeny in additional rounds of cutting with no further manipulation.

Permanent editing of the encoded target morphology without genomic editing reveals a new kind of epigenetics.

(See illustration on page 44.) In further experiments, we demonstrated that briefly reducing gap junction-mediated connectivity between adjacent cells in the bioelectric network that guides regeneration led worms to regenerate head and brain shapes appropriate to other worm species whose lineages split more than 100 million years ago.12 My group has developed the use of voltage-sensitive dyes to visualize the bioelectric pattern memory that guides gene expression and cell behavior toward morphogenetic outcomes.13 Meanwhile, my Allen Center colleagues are using synthetic artificial electric tissues made of human cells and computer models of ion channel activity to understand how electrical dynamics across groups of non-neural cells can set up the voltage patterns that control downstream gene expression, distribution of morphogen molecules, and cell behaviors to orchestrate morphogenesis.14,15 The emerging picture in this field is that anatomical software is highly modular—a key property that computer scientists exploit as subroutines and that most

likely contributes in large part to biological evolvability and evolutionary plasticity. A simple bioelectric state, whether produced endogenously during development or induced by an experimenter, triggers very complex redistributions of morphogens and gene expression cascades that are needed to build various anatomies. The information stored in the body’s bioelectric circuits can be permanently rewritten once we understand the dynamics of the biophysical circuits that make the critical morphological decisions. This permanent editing of the encoded target morphology without genomic editing reveals a new kind of epigenetics, information that is stored in a medium other than DNA sequences and chromatin.

Synthetic living machines and beyond Cells can clearly build structures that are different from their genomic-default anatomical outcomes. But are cells universal constructors? Could they make anything if only we knew how to motivate them to do it? The most recent advances in the new field at the intersection of developmental biology and computer science are driven by synthetic living machines known as biobots. Built from multiple interacting cell populations, these engineered machines have applications in disease modeling and drug development, and as sensors that detect and respond to biological signals. We recently tested the plasticity of cells by evolving in silico designs with specific movement and behavior capabilities and used this information to sculpt selforganized growth of aggregated Xenopus skin and muscle cells. 1 In a novel environment—in vitro, as opposed to inside a frog embryo—swarms of genetically normal cells were able to reimagine their multicellular form. With minimal sculpting post self-assembly, these cells form “Xenobots” with structures, movements, and other behaviors quite different from what might be expected if one simply sequenced their genome and identified them as wildtype X. laevis.

ORGANISMAL CONSTRUCTION

During morphogenesis, cells cooperate to reliably build anatomical structures. Many living systems remodel and regenerate tissues or organs despite considerable damage—that is, they progressively reduce deviations from specific target morphologies, and halt growth and remodeling when those morphologies are achieved. Evolution exploits three modalities to achieve such anatomical homeostasis: biochemical gradients, bioelectric circuits, and biophysical forces. These interact to enable the same large-scale form to arise despite significant perturbations.

 2  1  3

 1 BIOCHEMICAL GRADIENTS

© N.R. FULLER, SAYO-ART, LLC

The best-known modality concerns diffusible intracellular and extracellular signaling molecules. Gene-regulatory circuits and gradients of biochemicals control cell proliferation, differentiation, and migration.

 2 BIOELECTRIC CIRCUITS

The movement of ions across cell membranes, especially via voltage-gated ion channels and gap junctions, can establish bioelectric circuits that control largescale resting potential patterns within and among groups of cells. These bioelectric patterns implement long-range coordination, feedback, and memory dynamics across cell fields. They underlie modular morphogenetic decision-making about organ shape and spatial layout by regulating the dynamic redistribution of morphogens and the expression of genes.

 3 BIOMECHANICAL FORCES

Cytoskeletal, adhesion, and motor proteins inside and between cells generate physical forces that in turn control cell behavior. These forces result in largescale strain fields, which enable cell sheets to move and deform as a coherent unit, and thus execute the folds and bends that shape complex organs.

REWRITING THE PLANARIAN BODY PLAN Recent work from our group and others has demonstrated that anatomical pattern memories can be rewritten by physiological stimuli and maintained indefinitely without genomic editing. For example, the bioelectric circuit that normally determines head number and location in regenerating planaria can be triggered by brief alterations of ion channel or gap junction activity to alter the animal’s body plan. Due to the circuit’s pattern memory, the animals remain in this altered state indefinitely without further stimulation, despite their wildtype genomes. In other words, the pattern to which the cells build after damage can be changed, leading to a target morphology distinct from the genetic default.

 1

First, we soaked a planarian in voltage-sensitive fluorescent dye to observe the bioelectrical pattern across the entire tissue. We then cut the animal to see how this pattern changes in each fragment as it begins to regenerate.

 2 We then applied drugs or used RNA

interference to target ion channels or gap junctions in individual cells and thus change the pattern of depolarization/ hyperpolarization and cellular connectivity across the whole fragment.

 4

When we re-cut the two-headed planarian in plain water, long after the initial drug has left the tissue, the new anatomy persists in subsequent rounds of regeneration.

CREDIT © N.R. FULLER, LINE SAYO-ART, LLC

 3 As a result of the disruption of the body’s bioelectric circuits, the planarian regrows with two heads instead of one, or none at all.

These living creations are a powerful platform to assess and model the computations that these cell swarms use to determine what to build. Such insights will help us to understand evolvability of body forms, robustness, and the true relationship between genomes and anatomy, greatly potentiating the impact of genome editing tools and making genomics more predictive for large-scale phenotypes. Moreover, testing regimes of biochemical, biomechanical, and bioelectrical stimuli in these biobots will enable the discovery of optimal stimuli for use in regenerative therapies and bioengineered organ construction. Finally, learning to program highly competent individual builders (cells) toward group-level, goal-driven behaviors (complex anatomies) will significantly advance swarm robotics and help avoid catastrophes of unintended consequences during the inevitable deployment of large numbers of artificial agents with complex behaviors.16

Understanding how cells and tissues make real-time anatomical decisions is central to achieving regenerative outcomes too complex for us to manage directly.

In molecular biomedicine, we are still focused largely on manipulating the cellular hardware—the proteins that each cell can exploit. But evolution has ensured that cellular collectives use this versatile machinery to process information flexibly and implement a wide range of large-scale body shape outcomes. This is biology’s software: the memory, plasticity, and reprogrammability of morphogenetic control networks. The coming decades will be an extremely exciting time for multidisciplinary efforts in developmental physiology, robotics, and basal cognition to understand how individual cells merge together into a collective with global goals not belonging to any individual cell. This will drive the creation of new artificial intelligence platforms based not on copying brain architectures, but on the multiscale problem-solving capacities of cells and tissues. Conversely, the insights of cognitive neurobiology and computer science will give us a completely new window on the information processing and decision-making dynamics in cellular collectives that can very effectively be targeted for transformative regenerative therapies of complex organs. Michael Levin is the director of the Allen Discovery Center at Tufts University and Associate Faculty at Harvard University’s Wyss Institute. Email him at michael. [email protected]. M.L. thanks Allen Center Deputy Director Joshua Finkelstein for suggestions on the drafts of this story.

tissue via Notch signaling and regulation of proliferation,” J Neurosci, 35:4366–85, 2015. 5. V.P. Pai et al., “HCN2 channel-induced rescue of brain teratogenesis via local and long-range bioelectric repair,” Front Cell Neurosci, 14:136, 2020. 6. D.S. Adams et al., “Light-activation of the Archaerhodopsin H+-pump reverses agedependent loss of vertebrate regeneration: sparking system-level controls in vivo,” Biol Open, 2:306–13, 2013. 7. B.T. Chernet et al., “Use of genetically encoded, light-gated ion translocators to control tumorigenesis,” Oncotarget, 7:19575–88, 2016. 8. V.P. Pai et al., “Transmembrane voltage potential controls embryonic eye patterning in Xenopus laevis,” Development, 139:313–23, 2012. 9. A. Masotti et al., “Keppen-Lubinsky syndrome is caused by mutations in the inwardly rectifying K+ channel encoded by KCNJ6,” Am J Hum Genet, 96:295–300, 2015. 10. W.S. Beane et al., “A chemical genetics approach reveals H,K-ATPase-mediated membrane voltage is required for planarian head regeneration,” Cell Chem Biol, 18:77–89, 2011. 11. F. Durant et al., “The role of early bioelectric signals in the regeneration of planarian anterior/ posterior polarity,” Biophys J, 116:948–61, 2019. 12. M. Emmons-Bell et al., “Gap junctional blockade stochastically induces different species-specific head anatomies in genetically wild-type Girardia dorotocephala flatworms,” Int J Mol Sci, 16:27865-96, 2015. 13. D.S. Adams et al., “Bioelectric signalling via potassium channels: a mechanism for craniofacial dysmorphogenesis in KCNJ2associated Andersen-Tawil Syndrome,” J Physiol, 594:3245–70, 2016. 14. H.M. McNamara et al., “Bioelectrical domain walls in homogeneous tissues,” Nat Phys, 16:357–64, 2020. 15. H.M. McNamara et al., “Geometry-dependent arrhythmias in electrically excitable tissues,” Cell Syst, 7:359–70.e6, 2018. 16. M. Rubenstein et al., “ Programmable selfassembly in a thousand-robot swarm,” Science,

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345:795–99, 2014.

The emerging field of synthetic morphology emphasizes a conceptual point that has been embraced by computer scientists but thus far resisted by biologists: the hardware-software distinction. In the 1940s, to change a computer’s behavior, the operator had to literally move wires around—in other words, she had to directly alter the hardware. The information technology revolution resulted from the realization that certain kinds of hardware are reprogrammable: drastic changes in function could be made at the software level, by changing inputs, not the hardware itself.

References 1. S. Kriegman et al., “A scalable pipeline for designing reconfigurable organisms,” PNAS, 117:1853–59, 2020. 2. L.N. Vandenberg et al., “Normalized shape and location of perturbed craniofacial structures in the Xenopus tadpole reveal an innate ability to achieve correct morphology,” Dev Dyn, 241:863– 78, 2012. 3. K. Pinet et al., “Adaptive correction of craniofacial defects in pre-metamorphic Xenopus laevis tadpoles involves thyroid hormone–independent tissue remodeling,” Development, 146:dev175893, 2019. 4. V.P. Pai et al., “Endogenous gradients of resting potential instructively pattern embryonic neural

09. 2020 | T H E S C IE N T IST 4 5

EDITOR’S CHOICE PAPERS

The Literature Feather

Mosaic canary

Red siskin

GENETICS & GENOMICS

A Female Bird’s Muted Colors

OR

OR

THE PAPER

M. Gazda et al., “A genetic mechanism for sexual dichromatism in birds,” Science, 368:1270–74, 2020.

46 T H E SC I EN TIST | the-scientist.com

BCO2

European serin

BCO2

Common canary

DIALING DOWN THE GLITZ: In some finch species, BCO2, a gene that encodes a carotenoid-destroying

enzyme, is expressed in many female finch feathers but not in many male feathers. This generates dramatic sexual dichromatism that makes males dazzle while females look relatively drab. In common canaries, which are not sexually dichromatic, both males and females have little BCO2 expression in their feathers.

canary. They found one stretch of the mosaic genome that was associated with sexual dichromatism. They then looked for a region within that stretch that contained only siskin single nucleotide polymorphisms (SNPs) and found a sequence approximately 36 kilobases in length that contained three genes. The researchers measured the expression of those genes in regenerating tail feather follicles of mosaic canaries and found that only one, beta-carotene oxygenase 2 (BCO2), was expressed differently in females and males. When BCO2 is expressed, mosaic birds’ feathers are white because BCO2 degrades carotenoids. (In red siskins, color is bit more complex; they also possess a gene for melanin, which adds a gray tint.) Gazda next investigated whether other finch species also used BCO2 to generate sexual dichromatism, or if they’ve evolved different mechanisms. She found that BCO2 expression varied with the amount of carotenoids in feathers in the sexually dichromatic European serin (Serinus serinus) and in wild canaries from the Canary Islands (which show very slight sexual dichromatism), but not in the house finch (Haemorhous mexi-

canus), which must use another molecular mechanism. “The genes responsible for dichromatism are very exciting, and in this case it seems to be a common gene across finches (and possibly other birds!),” Pennsylvania State University evolutionary biologist David Toews, who was not involved in this study, writes in an email. Further experiments showed that it’s likely that an enhancer or promoter near the BCO2 gene turns it on, and Gazda suspects it is regulated by estrogen. “When the females get old [and make less of the hormone], they start to look a little bit more like males—they have more color,” she explains. Females without ovaries also look like males. Gazda says more experiments are needed to determine whether estrogen, or some other trigger, flips BCO2’s switch. “Identifying and characterizing the role that carotenoid processing genes [play], particularly this one, BCO2, has been elusive,” Toews writes, adding that the new study “has important consequences for our understanding about the evolution and genetics of animal coloration and signaling.” —Rachael Moeller Gorman

© KELLY FINAN

Male red siskins (Spinus cucullatus), a species of finch, flaunt orange-red bellies and backs, contrasting with their black heads and dark wing markings. The females, on the other hand, are mostly muted shades of grey (though pops of orange-red and black do appear on their bellies and wings). Such differences in coloration between the sexes, called sexual dichromatism, occur in many bird species, but their root cause has confounded scientists for years. Geneticist Miguel Carneiro of the Research Centre in Biodiversity and Genetic Resources (CIBIO) at the University of Porto in Portugal had previously discovered that a siskin gene called CYP2J19 encodes an enzyme that helps convert yellow carotenoid compounds from seeds in the birds’ diet into the red carotenoids found in their feathers. But he wanted to know why this happened only in males, so he and then–graduate student Małgorzata Gazda, together with Pedro Miguel Araújo of the nearby University of Coimbra, studied another, human-bred bird, the mosaic canary. These birds were created by breeding red siskins with common domestic canaries (Serinus canaria), which are not sexually dichromatic, and then backcrossing them with the common canaries and selecting for birds that are sexually dichromatic. Apart from their color and sexual dichromatism, mosaic canaries are nearly genetically identical to common canaries. Gazda sequenced the genomes of the mosaic canaries and compared them to the genomes of four non-dichromatic canary breeds and one slightly dichromatic wild

Follicle

WOUND RESIDENTS: Pathogenic bacteria such as Staphylococcus

IN THE BLOOD: RNA transcripts circulating in a mother’s blood yield

GENETICS & GENOMICS

DISEASE & MEDICINE

Wound-Healing Genes

Diagnosing Preeclampsia

THE PAPER

THE PAPER

C. Tipton et al., “Patient genetics is linked to chronic wound microbiome composition and healing,” PLOS Pathog, 16:e1008511, 2020.

S. Munchel et al., “Circulating transcripts in maternal blood reflect a molecular signature of early-onset preeclampsia,” Sci Transl Med, 12:eaaz0131, 2020.

© ISTOCK.COM, KSASS; © ISTOCK.COM, BUSRACAVUS

epidermidis, shown here, are more likely to emerge in wounds in which the diversity of other microbes is low.

How quickly scrapes, cuts, and gashes in the skin heal can vary greatly depending on a person’s body mass, age, and whether the individual suffers from certain chronic conditions such as diabetes. Genetics, a new study suggests, may also play a role, with variations in two specific genes lowering the diversity of a wound’s microbiome and lengthening healing time. Through a partnership with Southwest Regional Wound Care Center in Lubbock, Texas, geneticist Caleb Phillips at Texas Tech University and colleagues gained access to 85 patients’ DNA samples. Analyzing each person’s sample and comparing it to the diversity of bacteria in the patient’s infected wound, the team found that individuals with specific single nucleotide polymorphisms (SNPs) in TLN2, a gene involved in actin assembly, and ZNF521, which encodes a transcription factor, had lower overall microbial diversity in their wounds and were much more likely to suffer from Pseudomonas and Staphylococcus infections. Those patients’ skin injuries were also much slower to heal, suggesting that individuals with these specific TLN2 and ZNF521 mutations may be at higher risk of developing chronic wounds. Despite the “modest sample size,” Phillips says, the study offers a better understanding of what makes a patient vulnerable to chronic wounds. The SNPs the team described in TLN2 and ZNF521 could serve as biomarkers to identify patients at risk for slow wound recovery, he notes. The extent of the microbiome’s role in chronic wounds is “a really big question in the field of healing and repair,” notes Lindsay Kalan, a medical microbiologist and immunologist at the University of Wisconsin–Madison who was not involved in the study. While the paper’s results are “not immediately translatable” for patient care, she says, it is “definitely a step in the right direction.” —Lisa Winter

information about maternal, fetal, and placental health and might one day be used to diagnose preeclampsia.

Preeclampsia, a potentially fatal complication that affects roughly 5 percent of pregnancies worldwide, can only be diagnosed after the onset of symptoms such as high blood pressure, so treatment is always reactive. “The next really big need is better methods to diagnose or predict risk of pregnancy complications such as preeclampsia,” says Fiona Kaper, a senior director of scientific research at the biotech company Illumina. To identify possible biomarkers of the condition, Kaper and her colleagues drew blood from 40 pregnant women with earlyonset severe preeclampsia and 73 unaffected expecting mothers. Circulating in the blood of each mom-to-be is her own RNA, as well as transcripts from the placenta and the fetus. Studying these circulating RNAs (cRNAs), the team identified 30 maternal, fetal, or placental genes with altered expression patterns in women with preeclampsia compared with controls. A machine algorithm also identified 49 genes with altered expression, including 12 that overlapped with the earlier list, suspected of being linked to preeclampsia. To test the ability of the 49 suspect genes to predict preeclampsia, the researchers classified an independent cohort of two dozen women, half with early-onset preeclampsia and half without signs of the condition. The model predicted which women had preeclampsia with 85 percent to 89 percent accuracy. While large-scale, prospective studies are still needed, cRNA screening represents a step toward earlier preemptive diagnosis, says Kathryn Gray, an obstetrician at Brigham and Women’s Hospital who was not involved in the study. She notes that researchers have been doing something similar in detecting circulating tumor DNA for cancer screening. “It’s really exciting that we’re applying some of these . . . strategies that have been used in cancer to pregnancy. We’re always a bit behind in women’s health and pregnancy in applying the most cutting-edge technologies.” —Amanda Heidt 09. 2020 | T H E S C IE N T IST 47

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From Seeing to Analyzing

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Simplifying Progress

From Seeing to Analyzing Live-cell Imaging Before the advent of live-cell imaging, researchers relied on histology, observing fixed tissues adhered to slides to make biological conclusions. However, these cells were no longer living and did not fully recapitulate the complexities of living tissue and cells. In 1907 Julius Reis, a medical school professor at the Marey Institute in Paris recorded the first time-lapse film of sea urchin egg fertilization and embryonic development. Documenting a process that took fourteen hours to occur made it practical for Reis’s students to observe cell division and growth; the field of live-cell imaging was born.1 Live-cell imaging continued to evolve with the development of differential interference contrast (DIC) and other microscopy technologies, but the emergence of florescent dyes sparked the realization that researchers could use the technology to observe and track proteins.2-5 Fluorescent dye technology eventually led to flow cytometry, which offered, for the first time, a quantifiable and multiplexed measurement of fluorescently labeled cells.6 Flow cytometry technology quickly integrated into basic research and biotechnology, but it still required researchers to dissociate their cell populations to draw conclusions. The first fluorescence imaging whole plate reader made high throughput analysis of live-cell imaging possible by enabling researchers to take measurements from attached living cells.7

Live-cell Analysis Digital imaging microscopy expanded on these technologies by allowing multicolored, multimodal imaging. By transforming images into millions of tiny pixels, researchers began to interrogate visual images and collect data. This offered an unparalleled look at the dynamic nature of living cells. However, producing data required hours of human labor, focused on a small number of cells, and took weeks to months.2,8 In 1996, researchers developed the first fully automated high content screening (HCS) system, which removed the need for human labor. Automating image acquisition, processing, analyzing, and archiving enabled researchers to review multiple microplates with hundreds of cells in parallel, without human monitoring or interaction, in as little as one day. Unlike digital imaging microscopy, which created images as output data, HCS converted image data into digital data that researchers could use to make insightful conclusions about the dynamic nature of living cells.2 HCS systems revolutionized academic research and the pharmaceutical industry. Instead of screening one drug at a time for a singular biological effect selected by the investigator, researchers could screen hundreds of drugs and investigate their effects on a variety of cellular functions including the cell cycle, cell motility, apoptosis, toxicity, and more. Researchers moved from simply observing their cells to collecting functional information based on real-time screens with temporal and spatial information.2

Today, live-cell analysis systems deliver high-throughput multiplexed, multiparametric data from numerous multi-well plates, allowing researches to delve deeper into dynamic cellular activity.

Live-cell analysis systems enable researchers to peer into cells to observe and quantify dynamic cellular changes in real-time. These observations facilitate key discoveries that expand scientific knowledge and improve human health. Live-cell analysis systems lie at the intersection of live-cell imaging technology and high-content data analysis.

Digging Deeper

At the population level, researchers observe cellular kinetics, protein signaling, and cell-to-cell interactions in real-time to accurately measure population-level dynamics of primary cells from diseased patients and their response to possible treatments.

At the single-cell level, researchers delve deeper into the cell to observe subtle protein changes and capture rare cellular events that would otherwise go unnoticed in population-based observations.

More Possibil

The pharmaceutical and oncology industries analysis has played significant roles in the sci disease, cardiovascular research, stem cell re live-cell imaging as an invaluable tool for num

See references on the back.

The power behind HCS lies in its evolving software. First-generation software relied on readily available script-driven tools such as Metamorph and MATLAB and required specialized knowledge and training to operate. Second-generation software versions were designed specifically for HCS systems; these are still employed by some HCS systems today. These software versions allow fast and easy access to image analysis in an intuitive platform, making it suitable for novice users. While these software versions allow for some degree of flexibility in parameter and variable modification, their simplicity limits their capabilities. Rather than ready-to-go packages, third-generation software provides an application-agnostic interface in which the user defines the task and builds the analysis algorithm. These software versions allow users flexibility to design and execute multiplexed profile assays for a variety of cell responses in a single integrated analysis, allowing for a more efficient workflow than separate assays and analyses.9

Complex Culture

lities

s readily embraced live-cell analysis, which is well known for streamlining reliable candidate drug discovery. However, live-cell ientific advances of a number of other academic and research fields, including neuroscience, immunology, toxicology, infectious esearch, and inflammation. Studies analyzing cell health, cell function, and cell movement and morphology are quickly underscoring merous research fields.10

Cell Health & Kinetics Researchers rely on live-cell imaging systems to observe subtle changes in cellular kinetics, including growth, division, and health. Researchers also use live-cell imaging to automate and quantify the number of healthy cells based on morphology or in combination with a live/dead fluorescent indicator.

Cell Function Many live-cell imaging systems come equipped with fast imaging frame rates that, together with dispense and read capabilities, allow researchers to rapidly detect cellular responses that occur within milliseconds, such as those observed in calcium flux or cardiomyocyte contraction assays.

Cell Movement & Morphology Live-cell imaging platforms facilitate the observation and quantification of cell migration, a multistep process implicated in a number of biological and pathological processes, including embryonic development, tissue re-organization, angiogenesis, immune cell trafficking, chronic inflammation, wound healing, and tumor metastasis.11

3D Models Live-cell imaging and analysis systems facilitate the interrogation of biologically complex 3D structures, such as tumor spheroids and organoids. Imaging technology can solve the problem of identifying unique cell types in 3D cultures by identifying individual cell types through fluorescent labeling. Recent live-cell imaging systems come equipped with up to five fluorescent channels.12

Perfect Timing For some assays, it can be tricky to predict the optimal time for protein expression or other biological changes. Live-cell imaging takes away the guessing game by automatically and non-invasively capturing kinetic readouts from complex models that can be analyzed later to identify ideal timepoints for capturing and measuring biological events. For example, NOTCH signaling plays a key role in vascular tube formation, but is only active at later times in angiogenesis.12

Charting the Way in a New System Researchers are just beginning to explore cells derived from induced pluripotent stem cell (iPSC) differentiation. Cell types such as iPSC-derived neurons can take up to 55 days to mature through embryoid bodies, primitive neural stem cells, neural rosettes, and neural stem cells to become neurons. Using live-cell imaging and analysis systems, researches monitor cell health, morphology, and functional activity to make informed decisions about the best environmental components (growth factors, media, and surface coatings) needed to achieve the desired differentiation states for making key discoveries about development and cellular reprogramming.12

More Colors. More Insights. More Possibilities. See more information in every sample with the new Incucyte® SX5 featuring patent-pending optics. Do more with up to 5 colors specifically designed for live-cell analysis. www.sartorius.com/sx5 © 2020 Essen BioScience. All rights reserved. Incucyte and all Essen Bioscience products are registered trademarks and the property of Essen BioScience. Essen BioScience is a Sartorius Company.

Simplifying Progress

The Sartorius Group is a leading international partner of biopharmaceutical research and the industry. With innovative laboratory instruments and consumables, the Group’s Lab Products & Services Division concentrates on serving the needs of laboratories performing research and quality control at pharma and biopharma companies and those of academic research institutes. The Bioprocess Solutions Division with its broad product portfolio focusing on single-use solutions helps customers to manufacture biotech medications and vaccines safely and efficiently.

References H. Landecker, “Seeing things: from microcinematography to live cell imaging,” Nature, 6(10):707-709, 2009.

8. D. L. Farkas et al, “Multimode light microscopy and the dynamics of molecules, cells and tissues,” Annu Rev Physiol, 55:785–817, 1993.

2. D.L. Taylor, “Past, present, and future of high content screening and the field of cellomics,” Methods in Molecular Biology, Clifton, N.J. 356:3-18. 2007.

9. N. Thomas, “High-content screening: A decade of evolution,” Society for Biomolecular Sciences, 15(1): 2010.

3. A. H. Coons, M. M. Kaplan, “Localization of antigen in tissue cells. II. Improvements in a method for the detection of antigen by means of fluorescent antibody,” J Exper Med, 91:1– 13, 1950.

10. Sartorius, “Incucyte® SX5 live-cell analysis system,” Sartorius. https://www.essenbioscience.com/en/products/incucyte/incucyte-sx5-2020/?utm_expid=. HICTRiyGRJ6kYav9SkUMfA.1&utm_referrer=https%3A%2F%2Fwww.google.com%2F

4. A. S. Waggoner, “Dye indicators of membrane potential,” Ann Rev Biophys Bioeng, 8, 47–68, 1979

11. Sartorius, “Scratch wound migration and invasion assays for live-cell analysis,” Sartorius. https://www.essenbioscience.com/en/applications/live-cell-assays/scratch-wound-cellmigration-invasion/

1.

5. R. Haugland, “Intracellular ion indicators, in fluorescent and luminescent probes for biological activity,” (Mason, W. T., ed.), Academic, London, 34–43, 1993. 6. H. M. Shapiro, “Practical Flow Cytometry,” Fourth ed. Wiley-Liss, New York, 2003. 7.

K.S. Schroeder, B. D. Neagle, “FLIPR: a new instrument for accurate, high throughput optical sectioning,” J Biomol Screen, 1, 75–80. 1996.

12. Sartorius, “Interview: overcoming challenges of assays of complex cellular models with real-time live-cell analysis,” Sartorius, 2017. https://www.essenbioscience.com/en/about/decoding-cells-blog/interview-overcoming-challenges/

SCIENTIST TO WATCH

Ibrahim Cissé: Molecule Watcher Associate Professor, Department of Physics, MIT, Age: 37 BY JEF AKST

DENIS PAISTE, MIT MATERIALS RESEARCH LABORATORY

I

brahim Cissé spent hours as a child taking apart and trying to rebuild electronics. Other times, he would reenact NASA space shuttle liftoffs, tipping a chair backward onto the floor and then climbing in, his knees and face pointed skyward. He’d shake the chair to simulate shooting through Earth’s atmosphere as he watched Hollywood movies about space exploration. Captivated by the US space program as well as the idea of the American dream he saw portrayed on the big screen, the Niger native begged his parents to allow him to come to the US for college. They agreed, and with the help of a host family in North Carolina, he moved across the Atlantic at age 17. In 2002, Cissé transferred from community college to North Carolina Central University (NCCU), where he worked with physicist Kinney Kim—on a NASA-funded project, no less. “It was incredible,” Cissé says. But he didn’t realize that he could do research as a career until Nobel Laureate Carl Wieman and fellow physicist Sarah Gilbert, Wieman’s wife, visited NCCU during Cissé’s junior year and told him as much, Cissé recalls. “That was the first time someone had suggested to me the idea of doing . . . laboratory research for the rest of your life.” To prepare for graduate school, Cissé spent the summer of 2003 at Princeton University in the lab of physicist Paul Chaikin, now at New York University. Chaikin asked Cissé a seemingly simple question: If you poured ellipsoidal objects into a jar, how densely would they pack? The question had stumped previous students and many researchers, but Cissé had an idea. He filled a jar of M&Ms with paint, waited for it to dry, then removed the M&Ms by hand and counted the spots on each piece of candy where there was a missing circle of paint. Each spot indicated contact with another piece of candy, revealing that each candy has, on average, about 10 neighbors, he and his colleagues

reported in Science (303:990–93, 2004). That meant that ellipsoids packed more densely than perfect spheres, which have approximately six neighbors—a finding with implications for construction of ceramics, glass, and other materials. “It absolutely blew me away that this actually worked,” says Chaikin. “And it really was a discovery—it really was an influential paper that he was a coauthor on.” Cissé wrapped up his bachelor’s degree in 2004, then joined the lab of biophysicist Taekjip “TJ” Ha at the University of Illinois Urbana-Champaign. Ha had recently developed a variation of fluorescence resonance energy transfer (FRET) imaging that worked on the level of individual molecules. Cissé put a twist on the technique, housing the molecules of interest inside lipid vesicles, which kept them in close proximity so that they continued to interact despite not having strong affinity for each other. The method allowed Cissé to watch the assembly and disassembly dynamics of two mostly complementary strands of DNA with a single mismatch somewhere in the string of nine base pairs (Nat Struct Mol Biol, 19:623–27, 2012). After completing his PhD in 2009, Cissé accepted a position at École Normale Supérieure in Paris, where he developed a variation of a super-resolution microscopy technique called Photoactivated Localization Microscopy (PALM) to watch how molecules behaved over time in unfixed cells. The technique revealed that molecules of RNA polymerase II, a critical enzyme for transcription, cluster around genes that are being transcribed in cultured human cells (Science, 341:664–67, 2013). In 2014, Cissé started a lab at MIT, and following

up on the PALM work, he and his team reported that the length of time clusters hung around transcribed genes correlated with the number of mRNAs produced in mouse cells (eLife, 5:e13617, 2016). A later study showed that other molecules that regulate transcription clustered with RNA polymerases in mouse stem cells, and that some of the clusters lasted on the order of “tens of minutes,” Cissé says (Science, 361:412–15, 2018). “There’s a nice intellectual continuity to [his work],” says Ha, now at Johns Hopkins University, noting that Cissé has continued to focus on imaging the interactions of individual molecules. “I’m really, really impressed.” g

09. 202 0 | T H E S C IE N T IST 49

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MIKAEL KUBISTA, PHD Head Institute of Biotechnology Czech Academy of Sciences Founder and Chairman of the Board TATAA Biocenter

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CAREERS

When Science Meets Policy COVID-19 has laid bare some of the pitfalls of the relationship between scientific experts and policymakers—but some researchers say there are ways to make it better.

S

cience has taken center stage during the COVID-19 pandemic. Early on, as SARS-CoV-2 started spreading around the globe, many researchers pivoted to focus on studying the virus. At the same time, some scientists and science advisors—experts responsible for providing scientific information to policymakers—gained celebrity status as they calmly and cautiously updated the public on the rapidly evolving situation and lent their expertise to help governments make critical decisions, such as those relating to lockdowns and other transmissionslowing measures.

52 T H E SC I EN TIST | the-scientist.com

“Academia, in the case of COVID, has done an amazing job of trying to get as much information relevant to COVID gathered and distributed into the policymaking process as possible,” says Chris Tyler, the director of research and policy in University College London’s Department of Science, Technology, Engineering and Public Policy (STEaPP). But the pace at which COVID-related science has been conducted and disseminated during the pandemic has also revealed the challenges associated with translating fast-accumulating evidence for an audience not well versed in the

process of science. As research findings are speedily posted to preprint servers, preliminary results have made headlines in major news outlets, sometimes without the appropriate dose of scrutiny. Some politicians, such as Brazil’s President Jair Bolsonaro, have been quick to jump on premature findings, publicly touting the benefits of treatments such as hydroxychloroquine with minimal or no supporting evidence. Others have pointed to the flip-flopping of the current state of knowledge as a sign of scientists’ untrustworthiness or incompetence—as was seen, for exam-

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ple, in the backlash against Anthony Fauci, one of the US government’s top science advisors. Some comments from world leaders have been even more concerning. “For me, the most shocking thing I saw,” Tyler says, “was Donald Trump suggesting the injection of disinfectant as a way of treating COVID— that was an eye-popping, mind-boggling moment.” Still, Tyler notes that there are many countries in which the relationship between the scientific community and policymakers during the course of the pandemic has been “pretty impressive.” As an example, he points to Germany, where the government has both enlisted and heeded the advice of scientists across a range of disciplines, including epidemiology, virology, economics, public health, and the humanities. Researchers will likely be assessing the response to the pandemic for years to come. In the meantime, for scientists interested in getting involved in policymaking, there are lessons to be learned, as well some preliminary insights from the pandemic that may help to improve interactions between scientists and policymakers and thereby pave the way to better evidence-based policy.

Culture clash Even in the absence of a public-health emergency, there are several obstacles to the smooth implementation of scientific advice into policy. One is simply that scientists and policymakers are generally beholden to different incentive systems. “Classically, a scientist wants to understand something for the sake of understanding, because they have a passion toward that topic—so discovery is driven by the value of discovery,” says Kai Ruggeri, a professor of health policy and management at Columbia University. “Whereas the policymaker has a much more utilitarian approach. . . . They have to come up with interventions that produce the best outcomes for the most people.” Scientists and policymakers are operating on considerably different time-

scales, too. “Normally, research programs take months and years, whereas policy decisions take weeks and months, sometimes days,” Tyler says. “This discrepancy makes it much more difficult to get scientifically generated knowledge into the policymaking process.” Tyler adds that the two groups deal with uncertainty in very different ways: academics are comfortable with it, as measuring uncertainty is part of the scientific process, whereas policymakers tend to view it as something that can cloud what a “right” answer might be. This cultural mismatch has been particularly pronounced during the COVID19 pandemic. Even as scientists work at breakneck speeds, many crucial questions about COVID-19—such as how long immunity to the virus lasts, and how much of a role children play in the spread of infection—remain unresolved, and policy decisions have had to be addressed with limited evidence, with advice changing as new research emerges. “We have seen the messy side of science, [that] not all studies are equally well-done and that they build over time to contribute to the weight of knowledge,” says Karen Akerlof, a professor of environmental science and policy at George Mason University. “The short timeframes needed for COVID-19 decisions have run straight into the much longer timeframes needed for robust scientific conclusions.” Widespread mask use, for example, was initially discouraged by many politicians and public health officials due to concerns about a shortage of supplies for healthcare workers and limited data on whether mask use by the general public would help reduce the spread of the virus. At the time, there were few mask-wearing laws outside of East Asia, where such practices were commonplace long before the COVID19 pandemic began. Gradually, however, as studies began to provide evidence to support the use of face coverings as a means of stemming transmission, scientists and public health officials started to recommend their use. This shift led local, state, and federal officials

around the world to implement mandatory mask-wearing rules in certain public spaces. Some politicians, however, used this about-face in advice as a reason to criticize health experts. “We’re dealing with evidence that is changing very rapidly,” says Meghan Azad, a professor of pediatrics at the University of Manitoba. “I think there’s a risk of people perceiving that rapid evolution as science [being] a bad process, which is worrisome.” On the other hand, the spotlight the pandemic has put on scientists provides opportunities to educate the general public and policymakers about the scientific process, Azad adds. It’s important to help them understand that “it’s good that things are changing, because it means we’re paying attention to the new evidence as it comes out.”

Academia has done an amazing job of trying to get as much information relevant to COVID gathered and distributed into the policymaking process as possible. —Chris Tyler, University College London

Closing the gap Despite these challenges, science and policy experts say that there are both short- and long-term ways to improve the relationship between the two communities and to help policymakers arrive at decisions that are more evidence-based. Better tools, for one, could help close the gap. Earlier this year, Ruggeri brought together a group of people from a range of disciplines, including medicine, engineering, economics, and policy, to develop the Theoretical, Empirical, Applicable, Replicable, Impact (THEARI) rating system, a five-tiered framework for evaluating the robustness of scientific evidence in the con09. 202 0 | T H E S C IE N T IST 53

CAREERS

TALKING SCIENCE TO POLICYMAKERS For academics who have never engaged with policymakers, the thought of making contact may be daunting. Researchers with experience of these interactions share their tips for success. 1.

Do your homework. Policymakers usually have many different people vying for their time and attention. When you get a meeting, make sure you make the most of it. “Find out which issues related to your research are a priority for the policymaker and which decisions are on the horizon,” says Karen Akerlof, a professor of environmental science and policy at George Mason University.

2.

Get to the point, but don’t oversimplify. “I find policymakers tend to know a lot about the topics they work on, and when they don’t, they know what to ask about,” says Kai Ruggeri, a professor of health policy and management at Columbia University. “Finding a good balance in the communication goes a long way.”

3.

Keep in mind that policymakers’ expertise differs from that of scientists. “Park your ego at the door and treat policymakers and their staff with respect,” Akerlof says. “Recognize that the skills, knowledge, and culture that translate to success in policy may seem very different than those in academia.”

4.

Be persistent. “Don’t be discouraged if you don’t get a response immediately, or if promising communications don’t pan out,” says Meghan Azad, a professor of pediatrics at the University of Manitoba. “Policymakers are busy and their attention shifts rapidly. Meetings get cancelled. It’s not personal. Keep trying.”

5.

Remember that not all policymakers are politicians, and vice versa. Politicians are usually elected and are affiliated with a political party, and they may not always be directly involved in creating new policies. This is not the case for the vast majority of policymakers—most are career civil servants whose decisions impact the daily living of constituents, Ruggeri explains.

text of policy decisions. The ratings range from “theoretical” (the lowest level, where a scientifically viable idea has been proposed but not tested) to “impact” (the highest level, in which a concept has been successfully tested, replicated, applied, and validated in the real world). The team developed THEARI partly to establish a “common language” across scientific disciplines, which Ruggeri says would be particularly useful to policymakers evaluating evidence from a 5 4 T H E SC I EN TIST | the-scientist.com

field they may know little about. Ruggeri hopes to see the THEARI framework—or something like it—adopted by policymakers and policy advisors, and even by journals and preprint servers. “I don’t necessarily think [THEARI] will be used right away,” he says. “It’d be great if it was, but we . . . [developed] it as kind of a starting point.” Other approaches to improve the communication between scientists and policymakers may require more resources and time. According to Aker-

lof, one method could include providing better incentives for both parties to engage with each other—by offering increased funding for academics who take part in this kind of activity, for instance—and boosting opportunities for such interactions to happen. Akerlof points to the American Association for the Advancement of Science’s Science & Technology Policy Fellowships, which place scientists and engineers in various branches of the US government for a year, as an example of a way in which important ties between the two communities could be forged. “Many of those scientists either stay in government or continue to work in science policy in other organizations,” Akerlof says. “By understanding the language and culture of both the scientific and policy communities, they are able to bridge between them.” In Canada, such a program was established in 2018, when the Canadian Science Policy Center and Mona Nemer, Canada’s Chief Science Advisor, held the country’s first “Science Meets Parliament” event. The 28 scientists in attendance, including Azad, spent two days learning about effective communication and the policymaking process, and interacting with senators and members of parliament. “It was eye opening for me because I didn’t know how parliamentarians really live and work,” Azad says. “We hope it’ ll grow and involve more scientists and continue on an annual basis . . . and also happen at the provincial level.” There may also be insights from scientist-policymaker exchanges in other domains that experts can apply to the current pandemic. Maria Carmen Lemos, a social scientist focused on climate policy at the University of Michigan, says that one way to make those interactions more productive is by closing something she calls the “usability gap.” “The usability gap highlights the fact that one of the reasons that research fails to connect is because [scientists] only pay attention to the [science],” Lemos explains. “We are putting everything out

there in papers, in policy briefs, in reports, but rarely do we actually systematically and intentionally try to understand who is on the other side” receiving this information, and what they will do with it. The way to deal with this usability gap, according to Lemos, is for more scientists to consult the people who actually make, influence, and implement policy changes early on in the scientific process. Lemos and her team, for example, have engaged in this way with city officials, farmers, forest managers, tribal leaders, and others whose decision making would directly benefit from their work. “We help with organization and funding, and we also work with them very closely to produce climate information that is tailored for them, for the problems that they are trying to solve,” she adds. Azad applied this kind of approach in a study that involves assessing the effects of the pandemic on a cohort of children that her team has been following from

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The short timeframes needed for COVID-19 decisions have run straight into the much longer timeframes needed for robust scientific conclusions. —Karen Akerlof, George Mason University

infancy, starting in 2010. When she and her colleagues were putting together the proposal for the COVID-19 project this year, they reached out to public health decision makers across the Canadian provinces to find out what information would be most useful. “We have made sure to embed those decision makers in the project from the very beginning to ensure we’re asking the right questions, getting the most useful information, and getting it back to them in a very quick turnaround manner,” Azad says.

There will also likely be lessons to take away from the pandemic in the years to come, notes Noam Obermeister, a PhD student studying science policy at the University of Cambridge. These include insights from scientific advisors about how providing guidance to policymakers during COVID-19 compared to pre-pandemic times, and how scientists’ prominent role during the pandemic has affected how they are viewed by the public; efforts to collect this sort of information are already underway. “I don’t think scientists anticipated that much power and visibility, or that [they] would be in [public] saying science is complicated and uncertain,” Obermeister says. “I think what that does to the authority of science in the public eye is still to be determined.” g Diana Kwon is a freelance science journalist based in Berlin, Germany.

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VIRTUAL 2020 KEYNOTE PRESENTERS

Susan K. Gregurick, PhD Associate Director, Data Science (ADDS) and Director, Office of Data Science Strategy (ODSS), National Institutes of Health

Natalija Jovanovic, PhD Chief Digital Officer, Sanofi Pasteur

Bio-ITWorldExpo.com

READING FRAMES

Confronting Racism in Zoology A new book explores the history of scientists’ efforts to classify living things. BY DAVID BAINBRIDGE

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he death of George Floyd is having wide-ranging impacts, not just in the US but around the world. It has affected institutions and parts of society which previously believed themselves immune to, or at least removed from, racism. In my recent work on biological classification, I have been struck by how frequently science has been misused to reinforce existing prejudice. And alarmingly, although most overt racial science has been consigned to history, an inherent human obsession with biological classification has left a pervasive, ugly legacy: many people still believe some “races” to be more primitive than, or inferior to, others. I explore zoology’s history, warts and all, in my latest book, How Zoologists Organize Things. Humans seem driven to classify and organize, and the diversity of animal life around us serves as a perfect outlet for that urge. A particularly malign influence arose with the development in the Middle Ages of the scala naturae, a seemingly innocuous attempt to classify the natural (and theological) world according to a simple graduated system. In the early 14th century, the Majorcan philosopher Ramon Llull wrote Ladder of Ascent and Descent of the Mind, in which the elements of creation are allocated to steps on a staircase starting with minerals, then ascending through fire, plants, beasts, men, sky, and angels, ultimately overseen by God. Harmless as they may seem, the scalae implied a hierarchy of inherent value among natural things. These classifications of nature became progressively more complex, and with that complexity came a veneer of scientific authority, as in Charles Bonnet’s 1745 Notion of a Scale of Living Beings. As the 19th century started, these valueladen worldviews collided with new ways of

5 6 T H E SC I EN TIST | the-scientist.com

thinking in biology—especially the idea that humans could be analyzed and classified in the same way as other animals. To be sure, there have been several species of humans in Earth’s history, and science does classify these hominins based in part on physical characteristics, but early researchers took this idea too far by applying it to modern humans within the species Homo sapiens. For example, Charles White, in his disturbing 1799 book An Account of the Regular Gradation in Man and in Different Animals and Vegetables, ranks animals and people according to the verticality of their facial profile, from snipe and crocodiles to dogs and orangutans, and from “negroes,” “American savages,” and “Asiatics” to Europeans. During the first half of the 19th century, many zoologists’ minds turned toward the idea that animal types—and by extension, human types—could change and diverge over long periods of time. Yet this often meant the old hierarchy of nature simply transformed into a concept of evolutionary “progress” toward ever more perfect beings. And this carried the implication that some of those beings, human and animal, had been left behind in a primitive state. One of the strangest examples of this distortion of evolutionary theory was polygenism, an idea espoused by the pro-slavery scion of the American School of Ethnology, Josiah Nott. Although he disliked the concept of evolution, Nott, in his 1854 Types of Mankind, cherry-picked the ideas he needed to claim that the “human races” are distinct species with origins in different animal groups. Nott claimed the evidence showed that white men were justified in dominating Black men, whose attributes render them the perfect slaves. The respected German über-Darwinist Ernst Haeckel perpetuated the myth of evolutionary progress when he claimed influen-

White Lion Publishing, July 2020 tially that Judaism is an evolutionary intermediate between primitive paganism and advanced Christianity, and when he asserted that non-Europeans are “physiologically nearer to the mammals—apes and dogs— than to the civilized European. We must, therefore, assign a totally different value to their lives.” Zoology must fight hard to decontaminate itself from the value judgments and skewed arguments of the past. Already, we no longer speak of animals as primitive or advanced, and those concepts have become meaningless in the context of humans too. Additionally, the human species is no longer considered to be grouped into a number of discrete races, but rather an array of populations, each adapted to its ancestral geographical environment, yet blurring genetically and culturally into its neighbors. Humans have been the most widespread mammalian species for some time, so it is no surprise that we have ended up diverse. Yet it has taken a depressingly long time for scientists to state explicitly that this variation is messy and overlapping, and did not evolve to suit our prejudices. g David Bainbridge is a reproductive biologist and veterinary anatomist at the University of Cambridge. Read an excerpt of How Zoologists Organize Things at the-scientist.com.

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The Guide

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5 8 T H E SC I EN TIST | the-scientist.com

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FOUNDATIONS

Coronavirus Closeup, 1964 BY ASHLEY YEAGER

6 0 T H E SC I EN TIST | the-scientist.com

SAY CHEESE: In a 1964 paper containing the first published images of what would come to be known as a coronavirus, Donald Berry and his colleagues found that a pathogen that caused a deadly poultry disease (left) had widely spaced lollipop-shape spikes. By contrast, a particle of influenza A (right) had densely packed, rod-shape spikes.

particle. But the two types of viruses had different kinds of spikes. Influenza A’s were rod-shape and densely packed, while IBV’s were lollipop-shape, with bulbous growths at the ends, and they were much more spread out across the virus’s surface. IBV, Berry and his colleagues concluded in Virology in 1964, was not a form of the flu, as many had speculated (23:403–407). It was something else entirely. Early negative staining electron microscopy images of viruses such as those in the 1964 paper offered the “next big breakthrough” in virology after identifying whether families of viruses were RNA- or DNA-based, says Jeffrey Kahn, a University of Texas Southwestern Medical Center pediatrician and infectious disease researcher who has written about the history of coronavirus research. Such early images gave clues about the

true nature of viruses a decade before genetic sequencing became available. About a year after Berry’s publication, June Almeida of the St. Thomas’s Hospital Medical School in London identified another virus in the same family as IBV—one that infected humans. Looking through an electron microscope at virus particles isolated from samples from a boy suffering flulike symptoms in Surrey, England, she too noticed the virus had lollipop-like protrusions, which she’d seen before on IBV and other viral particles she’d studied. Drawing on the resemblance between the fringe of viral spikes and the halo of gas, or corona, surrounding the sun, Almeida and others, including Berry, wrote to Nature in 1968 to characterize IBV and other recently discovered pathogens like it as a new viral family called coronaviruses. g

IMAGES PUBLISHED IN VIROLOGY, “THE STRUCTURE OF INFECTIOUS BRONCHITIS VIRUS,” BY D.M. BERRY ET AL., © ELSEVIER, 1964

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ead baby chicks from farms began arriving by the dozens at the vet labs of North Dakota Agricultural College in Fargo in 1930. Chicken farmers also brought in their sick chicks, many of which were gasping for air. Others were listless and seemingly depressed, with drooping wings and emaciated bodies. In 20 years of lab and field research, vets Arthur Schalk and Merle Hawn had never seen a chicken disease quite like this one. It ripped through poultry farms in North Dakota and Minnesota that year, killing tens of thousands of baby birds. Based on necropsies of the dead birds, the vets ruled out laryngotracheitis, commonly called the croup, as a cause of death (JAVMA, 78:413–22, 1931). In that disease, lesions appear in the windpipe, but in these chicks, tissue damage was found farther down, deep in the lungs. The illness was a new one, and Schalk and Hawn named it infectious bronchitis of baby chicks. Alarmed at the impact on the poultry industry, other researchers began to investigate the disease, first identifying it as viral and naming the pathogen infectious bronchitis virus (IBV), and later revealing that its genetic code was RNA-based. It wasn’t until the 1960s that pathologists were able to get a look at the virus by staining the background of an electron microscope slide while leaving the actual specimen untouched, a technique called negative staining electron microscopy that was just emerging. Donald Berry of Glaxo Research, then located in a suburb of London, and his colleagues used the technique to capture fuzzy black-and-white images of an IBV particle and of an influenza A particle from a bird. As Berry and his colleagues stared at the images, the researchers could see spikes jutting out of each viral

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