IL-10 Receptor

Citation preview

IL-10 Receptor Rene de Waal Malefyt* Department of Molecular Biology, DNAX Research Institute, 901 California Avenue, Palo Alto, CA 94304-1104, USA * corresponding author tel: 650-496-1164, fax: 650-496-1200, e-mail: [email protected] DOI: 10.1006/rwcy.2000.14005.

SUMMARY IL-10 interacts with its tetrameric receptor complex consisting of two IL-10R and two IL-10R chains resulting in the phosphorylation and activation of JAK1 and TYK2 kinases, which in turn phosphorylate two tyrosine residues in the intracytoplasmic parts of the IL-10R chains that form docking sites for STAT3. Binding of STAT3 results in phosphorylation by JAK1 and TYK2 kinases, homo- or heterodimerization and translocation to the nucleus where it binds to the promoters of IL-10-responsive genes such as the Fc RI and activates transcription. All IL-10-mediated responses are dependent on activation of STAT3, but the anti-inflammatory actions of IL-10 require additional sequences in the distal intracytoplasmic part of the IL-10R chain.

BACKGROUND

Discovery IL-10 mediates its biological effects by interacting with specific cell surface receptors. The IL-10 receptor complex consists of at least two separate receptor chains, a ligand-binding IL-10R chain and a IL10R chain, which is essential for signal transduction. Radiolabeled IL-10 and FLAG epitope-tagged IL-10 were initially used to detect specific receptors on IL-10-responsive cells and to enrich for cells expressing high numbers of receptors (Tan et al., 1993). Expression cloning strategies led to the isolation of cDNAs encoding murine and human IL-10Rbinding proteins (Ho et al., 1993; Liu et al., 1994). Subsequently, an orphan class II cytokine receptor was identified as the IL-10R chain by functional

complementation studies in combination with the IL-10R chain (Kotenko et al., 1997).

Structure Both IL-10R and IL-10R belong to the class II cytokine receptor family, together with the receptors for IFN and IFN . Crystallization of human IL-10 or viral IL-10 to soluble extracellular IL-10R chains has indicated that the stoichiometry of the IL-10/sIL10R complex contains two IL-10 dimers binding to four shIL-10R monomers (Hoover et al., 1999), confirming results from previous gel filtration experiments (Tan et al., 1995). A model of IL-10 complexed with its soluble receptor at high resolution, based on topological similarity between IL-10 and IFN is also available (Zdanov et al., 1996).

Main activities and pathophysiological roles The main activity for the IL-10 receptor is to bind IL-10 and initiate the transduction of a signaling cascade, which leads to the modification of biological responses. There have been no specific pathophysiological roles described for the IL-10 receptor complex itself. The association of IL-10 with disease is described in the IL-10 chapter.

GENE

Accession numbers Mouse IL-10R: L12120 Human IL-10R: U00672, NM001558

1496 Rene de Waal Malefyt Mouse IL-10R : U53696 Human IL-10R : NM000628

Sequence

level. The first extracellular immunoglobulin-like binding domain of the hIL-10R lacks the intracellular disulfide bridge and the intracytoplasmic tail lacks one tyrosine residue as compared to the mIL-10R, although neither of these differences seem functionally significant.

See Figure 1.

Affinity for ligand(s) Chromosome location and linkages Mouse IL-10R: 9 Human IL-10R: 11q23.3 Mouse IL-10R : 16 Human IL-10R : 21q22.1±21q22.2 The structure of the IL-10R loci is conserved among human, mouse, and chicken but not in fish (Reboul et al., 1999).

PROTEIN

Accession numbers Mouse IL-10R: AAA16156.1 Human IL-10R: NP001549.1 Mouse IL-10R : AAC53062 Human IL-10R : NP000619

Description of protein The mIL-10R and hIL-10R polypeptides are composed of 576 and 578 amino acid residues respectively, whereas mIL-10R and hIL-10R chains consist of 349 and 325 amino acid residues respectively. IL-10R chains are expressed as 90±120 kDa proteins and IL-10R chains have a molecular weight of  40 kDa. IL-10R chains contain four (mouse) to six (human) N-linked glycosylation sites which are used. Positive cells express only a few hundred receptor complexes per cell.

Relevant homologies and species differences Mouse and human IL-10R chains are  60% identical and 73% similar. Mouse and human IL10R chains are 69% identical at the amino acid

Mouse and human IL-10 receptor complexes bind their ligand with high affinity (Kd  35±200 pM). Human IL-10 interacts with mIL-10R complex, but mouse IL-10 does not bind to the hIL-10R complex, which is consistent with the observed species specificities of these cytokines. The affinity and specific activity of human IL-10 is  10-fold lower on cells expressing mixed receptor complexes consisting of hIL-10R and mIL-10R chains as compared to mIL-10R complexes. EBV-derived viral IL-10 (vIL-10) bound human and mouse IL-10R transfectants with at least a 1000-fold lower affinity than their corresponding cellular ligand/receptor combinations. However, vIL-10 could activate cells through mIL10R or hIL-10R with a specific activity identical to the cellular cytokine depending on the cell type and in agreement with the limited biological spectrum of vIL-10 as compared to cellular IL-10. Recently, it has been shown that a single amino acid substitution at position 87 (I-A) is responsible for the reduced immunostimulatory activity of viral IL-10 (Ding et al., 2000). All responses to vIL-10 are dependent on the expression of the IL-10R as indicated by a blocking anti-hIL-10R monoclonal antibody (Liu et al., 1997). CmvIL-10, a viral homolog of IL-10 identified in human cytomegalovirus ORF UL111a, also binds to the hIL-10R complex and induces signal transduction events and biological activities (Kotenko et al., 2000). Interestingly, unlike EBV-derived vIL-10, which is 84% homologous to hIL-10, cmvIL-10 displays only 27% homology to hIL-10.

Cell types and tissues expressing the receptor IL-10R chains are expressed on cells of hematopoietic origin including T cells, B cells, monocytes, macrophages, dendritic cells, NK cells, mast cells, and various hematopoietic progenitors. In addition, tumor cells of hematopoietic origin, such as myeloma cells, B CLL, microglia, etc. have been described to express the IL-10R The IL-10R is present on epithelial cells from murine small and large intestine, on human

IL-10 Receptor 1497 Figure 1 Nucleotide sequences for the mouse and human IL-10R genes and the mouse and human IL-10R genes. Mouse IL-10Ra: 1 61 121 181 241 301 361 421 481 541 601 661 721 781 841 901 961 1021 1081 1141 1201 1261 1321 1381 1441 1501 1561 1621 1681 1741 1801 1861 1921 1981 2041 2101 2161 2221 2281 2341 2401 2461 2521 2581 2641 2701 2761 2821 2881 2941 3001 3061 3121 3181 3241 3301 3361 3421 3481

CCATTGTGCT ATGCCGCTGT CCTGAGCCTA TGAAGCCAGA CACCTACTAT CTGTAGAAAG CCGAAGCTAT GACCACCACT TCTGAAAGCA CCCTGCAGGG CCGGAAGTTC CACGGTCCCC AATTAACAAG CACTGTGACC CTGTCTGGTT CAAGAAGCCT TCACATCGTG GCATGGCAGC CCAATTCCTC TCCAGGGCTA CGGCTTACAC CCAGGATGAC GGATGCATCT CCAAGTCATG AGGCCCAGCA TGGGGTACAC GAAACAGGAG TCAACTGACC AAGTTGGAGG CCTGGATAGC AGAATGACAG TGGGAGGCTC CTAGCCAGGC GAGGTTGTGG GACAGAGACC GTAACCTCAG TGTCTGCGGA AGAAGGGCAG TGGGTTACTT ATTCAGAGCT TCTCTGCTAC ATTACCAGCA CCAGACGCTG TGGCCCAGGC AACAGGGTGC TCAGGGGACT GGTGCCCCTG GGAAGCCATC GGAGCCTAGG CCTCCCTATA AGGATGCTGA TCCAAATACA AGGTCATGAC CCCCCTGGAA AGGCGCAAGG TGACCTTGGG TTGGAAATTG AGCGGGCACA GTCAGGTGTT

GGAAAGCAGG GCGCCCAGGA GAATTCATTG TTTTTCCAGC GAAGTGGCCC GCTCAGGCAT GGCTACCGGG GAGACTCGCT ATGGACGGCA GATGAGTACG TCAGAACTAA ATAGGGGTGA GCAGAGTGGT AACCTGAGCA CTCCAGTGGT CACGACTTCT GACCTGGAGG ACCGACAGTG CTCCCTGGCT CAGGCCACCT TCCAGCATGG AGTGACGTTA GCCTTGGGCC GTGACATTCC GAATGCTTGG CTGCAGGATG TCTCAAGGGA GAAGAGTGGT TTTGCCCATA CTTGGCTCTA CGGCTAAGAG AGGAGTCAAA TCCACGGGGC CTAGTCTGCT TCCTCATGCC GGCTTTCTGG GGTGTTGGAG AGCATGAGCC GTGGCTGGGA GTGTGGCAAA CCATCTGCAG GTGGCCAAGC TCTGTCTGTG TTAGGACTTA TGGGCTGACA AAAATAATGA GGGTGATAGT TGTCCCCCTG GCCTTGTGCC CTGCGTTTGA GAGAAGCAGC GACTGTGTGA CCTAATCTGG CCTTTATTTA CTCTGGGTCT CAAGTCACGT AAATGTACCT GACGCCTCAA TTGTGGA

ACGCGCCGGC TGTTGTCGCG CATACGGGAC ACATCCTCCA TCAAACAGTA TGTCCTGTGA CCAGAGTCCG TCACAGTGGA TCATCTATGG AACAAGTCTT AGAATGCAAC GAAAGTTTTG CGGAGGAGCA TCTTAGTCAT ACATCCGGCA TCCCAGCCAA TTTTCCCAAA GCTTTGGCAG CCCACCCCCA GTGGGGACAA GGCCCGCCTG ACCTAGTCCA ATGTCTGTCT AGGGCTACCA ACGAAGAGAT ATTTGGCTTG TGGCTTCTGC CACTCCTGGG AACTTGACCC ACCTGGTCAC TTATTTGTAT GAAATATGTG AAGGAAAGGC GAGTGAGGGT TCAGGGCTGG ATGTGGTAAG TGGCTAGCCT AGGTTTAATT GATCTTGGGG AGGGACTGAG ACAGACATCT CATTACTCCC TTAGTACACT TGTCTGCTTT CCTCCGTGTT CTAGGTCATT TTAGGTCCTG GGGAGCTTCC AAGGGAGCTG GACCTGTCTT AAGGCTGCTG CGGGGTGAGG TACGAGAGCT TTTATTTGCT CTCAGGAGGT TTCCTCGTGG GACGTGCTCC ATGGAACCAC

CGGAGGCGTA TTTGCTCCCA AGAACTGCCA CTGGAAACCT CGGAAACTCA TCTCACAACG GGCAGTGGAC TGAAGTGATT GACAATCCAT CAAGGATCTC CAAGAGAGTG TGTCAAGGTG GTGTTTACTT ATCTATGCTG CCCGGGGAAG CCCTCTCTGC GGTGTCACTA TGGTAAACCA GATACAGGGG CACGGACAGT GAAGCAGCAG GAACTCTCCA CCTAGAACCT GAAACAGACC TCCCTTGACA GCCTCCACCA TCCACCAGGG TGTGGTTAGC TCTGGACTGT CCTGCCGTTG TCCAGCCATG GGTCCTTTTC CATCTTGATA CTGTAGATAC CTCCTACACT ACTGTAGGTC GCTACAGGAT TTGTCCTGTA TATACACCAC ACCCAGAATT TCATCTTTTT TGCTGCTCAC ACCCTTTAGG TGCTGCTAAT CAGCTGTGTG CAGAAGTCCC CAACCTCTGG ACCTCATGCC CTAGTCCCTG CAAATGGAGG ATCCCTGAGC CCAGCCATGA CCTTCTGGAA CACTTATTTA CTAGATTTGC AGCCTCAGTT ATCCCTAGGA AAGTGGTGTG

AAGGCCGGCT TTCCTCGTCA AGCCCTTCCT ATCCCAAACC ACCTGGAATG TTCACCCTGG AACAGTCAGT CTGACAGTGG CCCCCCAGGC CGAGTTTACA AAACAGGAAA CTGCCCCGCT ATCACGACGG CTATTCTGTG TTGCCTACAG CCAGAAACTC GAGCTGAGAG TCACTTCAGA ACTCTGGGAA GGGATCTGCC CTTGGATATA GGGCAGCCTA AAAGCCCCTG AGATGGAAGG GATGCCTTTG GCTCTGGCCG ACACCAAGTA TGTGAAGATC GGGGCAGCCC ATCTCCAGCC CCTGCTCCCC TGCAGACCTA CACGAGTGTC CAGCAGAGCT GGAAGGACCT TGAAGTCAGC AAAGGGAAGG GAGATGGTCC CCTGAATGAT TCTGTTCCTC ACTATGGCTG TGTTGTGACG TGGCCTTTGG CTCTAACTGC ACCTCCGACC TCATGCTGAA GTTGGAAGGA AGTGTTTCAG GGGTCTAGGG CAGTTTGCAG CCAGAGTTTC ACTTTGGCAT CTGGGCAAGC TTGAGGAAGC CTGCCCTGTT TTCCTGTCTG GTGCTGAGTC TGTTTTCATC

CCAGTGGACG CGATCTCCAG ATGTGTGGTT AGTCTGAGAG ACATCCATAT ATCTGTATCA ACTCCAACTG ATAGCGTGAC CCACGATAAC AGATTTCCAT CCTTCACCCT TGGAATCCCG AGCAGTATTT GAATCCTGGT TCCTGGTCTT CCGATGCCAT ACTCAGTCCT CTGAAGAGTC AAGAAGAGTC TGCAGGAGCC CCCATCAGGA AGTACACACA AGGAGAAAGA CAGAGGCAGC ATCCTGAACT CAGGTTATTT GACAGTGGAA TAAGCATAGA CTGGTGGCCT TGCAGGTAGA TCCCTGTACC CTGTGACCAG AGGTACATGA GAGCAGGATT GTGTTTGGGT TGAGCCTGGA CTCAAGAGAT CCAGCCAGGA CAGCCAGTCA TTGTGAGGTG TGTCCCCTGA TCAGACCAGA GCTTGAGCAC AGACCCAGAG AGCAGCTTCC TGTTAACCAA AGTGGACTAC AGATCTTGTG CTGGTCCCTG CCCCTAAGCA TCTGAAGCTT CCTGCCGAGA TCTTTGAGAC AGCGTGGCAC TCTAGCTGTG TATGCAAAGC CCACTGAGAA CTAATAAAAA

1498 Rene de Waal Malefyt Figure 1 (Continued) Human IL-10Ra: 1 61 121 181 241 301 361 421 481 541 601 661 721 781 841 901 961 1021 1081 1141 1201 1261 1321 1381 1441 1501 1561 1621 1681 1741 1801 1861 1921 1981 2041 2101 2161 2221 2281 2341 2401 2461 2521 2581 2641 2701 2761 2821 2881 2941 3001 3061 3121 3181 3241 3301 3361 3421 3481 3541 3601

AAAGAGCTGG GATGCTGCCG CGCTCATGGG CCACATCCTC GCTCCTGAGG CTATGACCTT GCGGGCTGTG GGATGAAGTG CGGGAAGATT CTTCAGTCAC CACACACAAG GTTCTGTGTC AGAGGAGTGC TGCCTTTGTC GCGGCGCCGA CATCAGCCAG CTTTTTGAAG CTTTGGCAGC TCACCCCCAG CAGTAGTGGC CCCCAGCACA CAGTGGCATT GGCCTTGGGC TGTGGCATTC AGGCTGCCTG CCTGGTTGAT TCCTCTAGAA TGAGGAATGG CTTTGCCCAT CTTTAACTCA CGGGCTGAGA GGGCCCTGGG GGCTGGCCCA ACTGAACTAG GACTCTGGCA CATAAAGGAT AGGCTGAAGT CAGCAGGAGG AGGGGAAAAA CAGGAAGCTT TAATGGATTC GAACCCATCC CAAATGGAGT TGCTGCCCCA AACTCAGCCC CAGGGTTGGA AGCCACTGGG CTAGCCACTT GGACCCTCCA CGTGGACTAC ATGGTGTCAT CTCCCCATAG AAGCTGTGAG TGGAGCATTC TGCCAAAGTA GGAGGATTCC GGAGAGGCAG CTCTGACTTT TCAGTTTCCT TGCTGAGGAA GTTTTTTATT

AGGCGCGCAG TGCCTCGTAG ACAGAGCTGC CACTGGACAC TATGGAATAG ACCGCAGTGA GACGGCAGCC ACTCTGACAG CAGCTACCCA TTCCGAGAGT AAAGTAAAAC CAGGTGAAAC ATCTCCCTCA CTGCTGCTCT AAGAAGCTAC CGTCCCTCCC GTGTCCCCAG ACCAAGCCAT GCTGACAGAA AGCAGCAATA GGGCCCACCT GACTTAGTTC CACCACAGTC CAGGGTTACC GAGGAAGAAT GAGGCAGGCT ATGACTCTGG TCACTCCTGG GACCTTGCCC GACCTGGTCA GGCTGCTTTT GCAGAAGTTA GCCAGGCTGC TGCAGGGTAT GAGCTGAGAA TATTTGCTCA CAGCTCAGAC AAGGGCTGTA GGAGGATATG GTCACTGGAA ACTGAGGGGA CTCTGGTGGG AGCATCCCCC CCATCTTGCT TTTGGGCGGC GGCCTGTGCT CACTGGGCTG GTCAGAGGGC TGTTTGCTGG CAAGCTGGCT GCCAAGACAG GCCATTTGGA GGGACAGGCC TGAAAACAGA CTCTTAGGTG TCAGGGTTCC CATTGCACAG GCTGTTTCCA CATCTGCAGA ATGGGTATGA CCAATAAATT

GCCGGCTCCG TGCTGCTGGC CCAGCCCTCC CCATCCCAAA AGTCCTGGAA CCTTGGACCT GGCACTCCAA TTGGCAGTGT GGCCCAAGAT ATGAGATTGC ATGAAAACTT CATCTGTCGC CCAGGCAGTA CCGGAGCCCT CCAGTGTCCT CAGAGACCCA AGCTGAAGAA CCCTGCAGAC CGCTGGGAAA GCACAGACAG GGGAGCAACA AAAACTCTGA CCCCGGAGCC TGAGGCAGAC CGCCCTTGAC TGCATCCACC CTTCCTCAGG CCTTGAGCAG CTCTAGGCTG CCCTGCCCCT GATTTTAGCC GGCACGAGGC AGGGCTGGTC GTGGGTGGCA GGGCAGGGAC GGGGAACCAT CCAGACCTCC GGAATGGAAG ATGGTCACAT GATCTTAAGG GACAAAGGGA TACCTCTGGC TGGGGCACTT GACAACTTCC CTCTGGGCTT TGTGTTTGCT GGGTCCCTGC CTCAATCTCC GTATTAGCCA TGTTTCTTAT TATCAGACAC CTCTGCCTTC TGTGCGTGCC TATTCTGGCC CCAGTCTGGT CTTGAAAGCT TGAAAGAATT GTGGTATGAC ATAATGACTG ATGTGCCTTG GTCAAGACCA

CTCCGGCCCC GGCGCTCCTC GTCTGTGTGG TCAGTCTGAA CTCCATCTCC GTACCACAGC CTGGACCGTC GAACCTAGAG GGCCCCCGCG CATTCGCAAG CAGCCTCCTA TTCCCGAAGT TTTCACCGTG CGCCTACTGC GCTCTTCAAG AGACACCATC CTTGGACCTG TGAAGAGCCC CGGGGAGCCC CGGGATCTGC GGTGGGGAGC GGGCCGGGCT TGAGGTGCCT CAGATGTGCT AGATGGCCTT AGCCCTGGCC GGCCCCAACG CTGCAGTGAC TGTGGCAGCC CATCTCTAGC ATGCCTGCTC AGTCTGGGCA AGGGTGTCTG CTGACCTGTT CTTCTCCCTC GGGGCTTTCT CTGCTTAGGC CTTCAGGGCC GGGGAACCTC TATATATTTT GCCGAGACCC ACCCATCTGC GCTGAGGCCA AGAGAAGCCA GGGCACCAGC GCTAATGTCC CTTGTTGGTG CATCTGTGAA AGCTGGTCCT GCCAGAGGCT AGCCCCAGAA AAACAAAGGC ATCCAGAGTC CAGGGAATCC AACTGAACTC TTATTTATTT CTGGATATCT CTTGGAGAAG ACTTGTCTAA AACACAAAGC CA

GGACGATGCG AGCCTCCGTC TTTGAAGCAG AGTACCTGCT AACTGTAGCC AATGGCTACC ACCAACACCC ATCCACAATG AATGACACAT GTGCCGGGAA ACCTCTGGAG AACAAGGGGA ACCAACGTCA CTGGCCCTCC AAGCCCAGCC CACCCGCTTG CACGGCAGCA CAGTTCCTCC CCTGTGCTGG CTGCAGGAGC AACAGCAGGG GGGGACACAC GGGGAAGAAG GAAGAGAAGG GGCCCCAAAT AAGGGCTATT GGACAGTGGA CTGGGAATAT CCAGGTGGTC CTGCAGTCAA CTCTGCCTGG CTTTTCTGCA GGGCAGGAGG CTGTTGACTG CTAGGAACTC GGAGTTGTGG CACTCGAGCA TTGCTGCTGG CCCTCATCGG CTGGACACTC TGGATGGGGC AAATATCTCC AGCCACTCAC TGGTTTTTTG TCATGCCAGC AGCTACAGAC TTCAGCTGTG ATAAGGACTC GGGAGAATGC AACAGATCCA GGGGGCATTA AGTTCAGTCC ATCTCAGCCC AGCCATGACC CCTCTGGAGG ATTTTGTTCA CAGGAGCCCC TCACTTATCC TTCATAGGGA TCTGTCAATA

GCGCGCCCAG TTGGCTCAGA AATTTTTCCA ATGAAGTGGC AGACCCTGTC GGGCCAGAGT GCTTCTCTGT GCTTCATCCT ATGAAAGCAT ACTTCACGTT AAGTGGGAGA TGTGGTCTAA TCATCTTCTT AGCTGTATGT CCTTCATCTT ATGAGGAGGC CAGACAGTGG TCCCTGACCC GGGACAGCTG CCAGCCTGAG GCCAGGATGA AGGGTGGCTC ACCCAGCTGC CAACCAAGAC TCGGGAGATG TGAAACAGGA ACCAGCCCAC CTGACTGGAG TCCTGGGCAG GTGAGTGACT ACCAGGAGGA AGTCCACTGG AGGCCAACTC GGGCCCTGCA TTTCCTGTAT TGAGGCCACC TCAGAGCTTC GGTCATTTTT GCCTCTGGGG AAACACATCA TTCCAGCTCA CTCTCTCCAA ATCCTCACTT TATTGGTCAT CCCAGAGGGT CCAGAGGATA TGATTTTGGA CACCTTTAGG AGATACTGTC ATGGGAGTCC TGGGCCCTGC ACAGGCATGG TGCCTTTCTC CCCACCCCTC CAGGCTTGAG TTTATTTATT GAAATTCTAG TCTTGGAGCC TGTGAGGTTC AGTGATACAT

IL-10 Receptor 1499 Figure 1 (Continued) Mouse IL-10Rb: 1 61 121 181 241 301 361 421 481 541 601 661 721 781 841 901 961 1021 1081 1141 1201 1261 1321 1381 1441 1501 1561 1621

ACATGGCCCC TGATTCCACC GGGAGGTACC GGTCTTTCCA CTAAATACGG GGGTCAATGT TAGAATCCCT CTGAGACGTG AAAATGGGAC ACCTGGAGCC GAACAGGAGA CCTGGATTGT GCTGCTTTGT CGTCTCTTCC TGTTCTCCTT AAGAGTCTGA ATCCAGGGCC CCAAACTGCT TCTAGAACTT TACTTGGGAA TGACTAGAGC ACCAGGGATG ACAGTCATCT GAGAGAGAGA TCACTAAAAC GGGCATGGAA GTTGGCCCCA TTCCTTACCT

GTGCGTGGCG CCCTGAGAAG TGCTTTCCCC AGATCACTGC AGACTACACT CACCTTCTGC TGCTGAGTCT GACCTTGAAG TAATGAGAAG GTGGACAACT GTGGAGTGAA GGCCATCATC CGTGCTGTGG ACAGCACCTG CCCTCCCCCC AGGCAGCAAG TCGGGAGCTG CACGTCGACC CCTGATGCTG TTTGCTGCCA TCTTACTTGG GGGGCACCTA TGAACTATAC GAGAACTTAT AAGGATCACA ACACTATGAA TGCGGGACAG GTGTGTTTTG

GGCTGGCTGG GTCAGAATGA AAAACGAACC AAGCGCACTG GTGAGAGTCA CCCGTGGAAG TTACACCTGC AACATTTATG TTTCAAGTTG TACTGCATTC CCCATCTGTG CTCATAGTCT CTCATTTATA AAGGAGTTTC GAGGAGGCCG CAGAGTCCTG GAGTCCAAGG TCAGAAGTAT CACTGGTACA TATAAAGACT CACACATGAA CCTATAAGCC AGGGAGTTCA ATATTTTATG TTTAACTTGT ATTATAAGAA ACATGAACAT TATTAATATT

GTGGCTTCCT ATTCAGTTAA TGACTTTCAC CCTCGACTCA GGGCTGAATT ACACCATCAT GTTTCTCAGC ACTCATGGGC TGTCTCCGTA AAGTTCAAGG AACGGACAGG CCGTCCTGGT AGAAGACCAA TGGGCCACCC AGGTGTTCGA AAGACAACTG ATGAAGCTCC GACCAGAGAG CACAACCAAA AATAATTTAG GTCCTAGCTT CAGCACTTTG AGGCCAAGCT GCCACTGAAT GACAAACAAA TGCCTATAGA TTTGGATTCC AGTGTTCTGT

TCTGGTGCCA TTTCAAGAAC AGCTCAGTAT GTGCGACTTC GGCGGATGAA TGGACCTCCT CCCACAAATT TTACAGAGTG CGACTCTGAG GTTTCTTCTC CAATGACGAA GGTCTTCCTC GCATACCTTC CCATCACAGC CAAACTAAGC TGCCTCAGAG CTCACCTCCA CCACCTGAAA GAGCTAGGTT GGACTGAGGG CGATCCCCAA GAGGTAGAGG GGACTAGAGA GTAATTTGAG AATATTTTAA CCACCCGCAT CAAGGAGCAA AAATATTCTA

GCTCTAGGAA ATTCTACAGT GAAAGTTACA TCTCATCTTT CATTCGGAGT GAGATGCAGA GAGAATGAGC CAATACTGGA GTCCTCCGGA GACCAGAACA ATAACCCCTT TTCCTCCTGG CGTTCTGGGA ACGTTTCTGC ATCATCAGCG CCCCCGTCTG CACGATGACC AAACTCCAAA TTAAACACTC TGTAGCTCAG CACCATATAA CAGGAGGATC CCCTGTCTAA CCCTTTGTGC ATGGGGGGGG CTCAAAAGTG AGAGATTTCC

Human IL-10Rb: 1 61 121 181 241 301 361 421 481 541 601 661 721 781

ATGGCGTGGA GTACCACCTC GAGTCACCTG ATATTCCAAG AAGTATGGTG GTAAACATCA GAAGTACTTG GAAACTTGGA AACGGTACTG CTGGAGCCAT GCTGGGGAAT TGGATGGTGG TGCTTCTCCT TCTCTTCCAC

GTCTTGGGAG CCGAAAATGT CTTTTGCCAA ATAAATGCAT ACCACACCTT CCTTCTGTCC ATGATTCTTT CTATGAAGAA ATGAAAAGTT GGACAACTTA GGAGTGAGCC CCGTCATCCT TGCTGTGGTG AGCACCTGAA

CTGGCTGGGT CAGAATGAAT AGGGAACCTG GAATACTACC GAGAGTCAGG TGTGGATGAC ACATATGCGT TGTGTATAAC TCAAATTACT TTGTGTTCAA TGTCTGTGAG CATGGCCTCG CGTTTACAAG AGAGGTAGGT

epidermal cells and keratinocytes and can be induced on fibroblasts (Michel et al., 1997; Denning et al., 2000). IL-10R chain seems to be expressed ubiquitously.

Regulation of receptor expression IL-10R expression is induced in fibroblasts by LPS activation (Weber-Nordt et al., 1994). In contrast, the

GGCTGCCTGC TCTGTTAATT ACTTTCACAG TTGACGGAAT GCTGAATTTG ACCATTATTG TTCTTAGCCC TCATGGACTT CCCCAGTATG GTTCGAGGGT CAAACAACCC GTCTTCATGG AAGACAAAGT AGGATGGAGT

TGGTGTCAGC TCAAGAACAT CTCAGTACCT GTGATTTCTC CAGATGAGCA GACCCCCTGG CTAAAATTGA ATAATGTGCA ACTTTGAGGT TTCTTCCTGA ATGACGAAAC TCTGCCTGGC ACGCCTTCTC GA

ATTGGGAATG TCTACAGTGG AAGTTATAGG AAGTCTTTCC TTCAGACTGG AATGCAAGTA GAATGAATAC ATACTGGAAA CCTCAGAAAC TCGGAACAAA GGTCCCCTCC ACTCCTCGGC CCCTAGGAAT

expression of IL-10R mRNA by human T cell clones was downregulated following activation (Liu et al., 1994).

Release of soluble receptors No data has been published on the production and release of soluble IL-10 receptors in normal or disease states.

1500 Rene de Waal Malefyt

SIGNAL TRANSDUCTION

Associated or intrinsic kinases Neither the IL-10R nor the IL-10R chain possesses an intrinsic kinase activity. However, IL-10 treatment of cells induces phosphorylation of the IL-10Rassociated JAK1 and the IL-10R -associated TYK2 kinases (Finbloom and Winestock, 1995; Ho et al., 1995).

Cytoplasmic signaling cascades Phosphorylation of JAK1 (Janus kinase 1) and TYK2 kinases by IL-10 binding to the receptor complex activates their kinase activity and leads to phosphorylation of two tyrosine residues, which are part of the cytokine receptor box 3 motif, (Y446 and Y496 in hIL-10R and Y427 and Y477 in mIL-10R) in the intracytoplasmic tail of the IL-10R chain. Phosphorylation of at least one tyrosine residue is sufficient for a biological response. These phosphorylated tyrosine residues then serve as docking sites for the latent transcription factor STAT3 (signal transducer and activator of transcription 3) which in turn becomes phosphorylated by the activated JAK1 and TYK2 kinases. Phosphorylated STAT molecules subsequently form homo- or heterodimers and translocate from the cytoplasm to the nucleus, where they bind with high affinity to STAT-binding elements (SBE) in the promoters of IL-10-responsive genes (Lai et al., 1996; Wehinger et al., 1996). IL-10 can also activate STAT5 and/or only STAT1 DNA-binding activity in some cell types (Weber-Nordt et al., 1996a; Zocchia et al., 1997). Whereas STAT3 is directly recruited to the IL-10R chain, activation of STAT1 and STAT5 may occur through other mechanisms (Weber-Nordt et al., 1996b). The two membranedistal tyrosine molecules in the intracytoplasmic tail of the IL-10R chain and STAT3 are essential for the antiproliferative and developmental actions of IL-10, but an additonal C-terminal sequence which contains at least one functionally critical serine residue is required for the anti-inflammatory effects of IL-10 such as inhibition of TNF production by monocytes/macrophages (O'Farrell et al., 1998; Riley et al., 1999). Another membrane-proximal region of the IL10R chain has been implicated as a functional domain involved in regative regulation (Ho et al., 1995). IL-10 antagonizes the induction of some IFNor IFN -inducible genes on human monocytes by preventing the IFN-induced phosphorylation of

STAT1 (Ito et al., 1999). This may be mediated by the rapid STAT3-dependent induction of SOCS3 (suppressor of cytokine signaling 3). SOCS proteins are members of a family of molecules that interfere with JAK/STAT signal transduction pathways. Alternatively, differences in DNA-binding activity and composition of IL-10-induced STAT1 and IFNinduced STAT1 complexes may play a role in the differential responses to IL-10 and IFN (Yamaoka et al., 1999). Interestingly, in contrast to IFN , IL-10 failed to upregulate Fc RI expression and GRR (IFN response region)- or SIE (serum-inducible element)binding activity in human neutrophils, despite the presence of a functional IL-10R complex, and STAT1 and STAT3 expression by these cells (Bovolenta et al., 1998). IL-10 did induce SOCS3 expression in neutrophils, indicating that activation of STAT1 or STAT3 phosphorylation is not required for SOCS3 induction in these cells as it is in monocytes (Cassatella et al., 1999). Other signaling pathways that have been implicated in biological responses to IL-10 include NFB, phosphatidylinositol 3-kinase, p70 S6 kinase, and MAP kinase cascades, but none of these have been directly linked to molecules that interact with the IL-10 receptor (De Waal Malefyt and Moore, 1998).

DOWNSTREAM GENE ACTIVATION

Transcription factors activated IL-10 activates STAT1, STAT3, and in some cell types STAT5 DNA-binding activity and transcriptional activation (Finbloom and Winestock, 1995; Ho et al., 1995; Lai et al., 1996; Weber Nordt et al., 1996a; Wehinger et al., 1996).

Genes induced IL-10 induces expression of Fc RI, TIMP-1 (tissue inhibitor of metalloproteinases 1), MCP-1, Ilinck (IL10-induced chemokine), CCR5 on monocytes, and enhances IL-1Ra and soluble p55 and p75 TNFR. On mouse mast cells it induces expression of mast cell-specific proteases mMCP-1 and mMCP-2, whereas on mouse B cells it enhanced expression of MHC class II. On human B cells it enhanced expression of Bcl-2 and the high-affinity IL-2R. A protein homologous to SNAP23 (synaptosomal-associated protein of 23 kDa) was induced by IL-10 in OTT1 cells (Morikawa et al., 1998). IL-10 inhibited the

IL-10 Receptor 1501 proliferation of bone marrow-derived macrophages and of J774 cells by STAT3-dependent induction of the cyclin-dependent kinase inhibitor p19INK4D, which acts on the interactions between cdk's 4 and 6 and D cyclins, and the STAT3-independent induction of p21CIP1, which has been shown to inhibit cyclins A or E-associated cdk-2 activity.

birth and developed normally until 12 weeks of age, at which time a majority developed a chronic colitis and splenomegaly, reminiscent of the pathology and phenotype of IL-10 knockout mice (Spencer et al., 1998). Cells from IL-10R ÿ/ÿ mice responded normally to type I and type II interferons, but did not respond to IL-10.

Promoter regions involved

Human abnormalities

IL-10-activated STAT1, STAT3, and STAT5 are able to bind to the GRR (IFN response region) of the Fc RI gene, the SIE (serum-inducible element) of the c-fos promoter and the PRL-STAT (prolactin STAT) consensus sequence of the -casein gene (Wehinger et al., 1996). In addition, STAT3 bound the IL-6 response element in hepatoma cells transfected with the IL-10R (Lai et al., 1996) and could enhance the expression of hsp90 (heat shock protein) and hsp90 promoters in these cells and in peripheral blood mononuclear cells (Ripley, 1999). Finally, IL-10 activated two STAT3-binding sites in the proximal p19INK4D promoter (O'Farrell et al., 2000). IL-10 downregulated IFN -induced ICAM1 transcription in human monocytes by preventing IFN induced binding activity at the NFB site of the TNF-responsive NFB/CEBP composite element in the ICAM1 promoter (Song et al., 1997).

No human abnormalities related to the IL-10R complex have been described to date.

BIOLOGICAL CONSEQUENCES OF ACTIVATING OR INHIBITING RECEPTOR AND PATHOPHYSIOLOGY

Unique biological effects of activating the receptors IL-10 binding to its receptor complex induces the modulation of a wide variety of immunological and biological responses as described in the IL-10 chapter. However, the most prominent activity of IL-10 is probably its immunosuppressive and anti-inflammatory actions on monocytes (De Waal Malefyt and Moore, 1998).

Phenotypes of receptor knockouts and receptor overexpression mice An IL-10R chain knockout mouse has been described. These mice did not show abnormalities at

THERAPEUTIC UTILITY

Effect of treatment with soluble receptor domain ShIL-10R are able to neutralize the effects of IL-10 in proliferation and differentation assays (Tan et al., 1995).

Effects of inhibitors (antibodies) to receptors Anti-IL-10R antibodies that neutralize the biological activity of mIL-10 or hIL-10 have been described (Ho et al., 1995; Liu et al., 1998). Although in vivo studies in mice are ongoing, it can be predicted that anti-IL10R monoclonal antibodies, like anti-IL-10 monoclonal antibodies, are able to enhance acquired cellular immune reponses against intracellular pathogens, bacteria, and tumor cells (De Waal Malefyt and Moore, 1998).

References Bovolenta, C., Gasperini, S., McDonald, P. P., and Cassatella, M. A. (1998). High affinity receptor for IgG (Fc gamma RI/CD64) gene and STAT protein binding to the IFN-gamma response region (GRR) are regulated differentially in human neutrophils and monocytes by IL-10. J. Immunol. 160, 911±919. Cassatella, M. A., Gasperini, S., Bovolenta, C., Calzetti, F., Vollebregt, M., Scapini, P., Marchi, M., Suzuki, R., Suzuki, A., and Yoshimura, A. (1999). Interleukin-10 (IL-10) selectively enhances CIS3/SOCS3 mRNA expression in human neutrophils: evidence for an IL-10-induced pathway that is independent of STAT protein activation. Blood 94, 2880±2889. De Waal Malefyt, R., and Moore, K. W. (1998). In ``The Cytokine Handbook, 3rd edn'' (ed A. Thomson), Interleukin-10, pp. 333± 364. Academic Press, London.

1502 Rene de Waal Malefyt Denning, T. L., Campbell, N. A., Song, F., Garofalo, R. P., Klimpel, G. R., Reyes, V. E., and Ernst, P. B. (2000). Expression of IL-10 receptors on epithelial cells from the murine small and large intestine. Int. Immunol. 12, 133±139. Ding, Y., Qin, L., Kotenko, S. V., Pestka, S., and Bromberg, J. S. (2000). A single amino acid determines the immunostimulatory activity of interleukin 10. J. Exp. Med. 191, 213±224. Finbloom, D. S., and Winestock, K. D. (1995). IL-10 induces the tyrosine phosphorylation of tyk2 and Jak1 and the differential assembly of STAT1 alpha and STAT3 complexes in human T cells and monocytes. J Immunol 155, 1079±1090. Ho, A. S., Liu, Y., Khan, T. A., Hsu, D. H., Bazan, J. F., and Moore, K. W. (1993). A receptor for interleukin 10 is related to interferon receptors. Proc. Natl Acad. Sci. USA 90, 11267± 11271. Ho, A. S., Wei, S. H., Mui, A. L., Miyajima, A., and Moore, K. W. (1995). Functional regions of the mouse interleukin-10 receptor cytoplasmic domain. Mol. Cell Biol. 15, 5043±5053. Hoover, D. M., Schalk-Hihi, C., Chou, C. C., Menon, S., Wlodawer, A., and Zdanov, A. (1999). Purification of receptor complexes of interleukin-10 stoichiometry and the importance of deglycosylation in their crystallization. Eur. J. Biochem. 262, 134±141. Ito, S., Ansari, P., Sakatsume, M, Dickensheets, H., Vazquez, N., Donnelly, R. P., Larner, A. C., and Finbloom, D. S. (1999). Interleukin-10 inhibits expression of both interferon alpha- and interferon gamma-induced genes by suppressing tyrosine phosphorylation of STAT1. Blood 93, 1456±1463. Kotenko, S. V., Krause, C. D., Izotova, L. S., Pollack, B. P., Wu, W., and Pestka, S. (1997). Identification and functional characterization of a second chain of the interleukin-10 receptor complex. EMBO J. 16, 5894±5903. Kotenko, S. V., Saccani, S., Izotova, L. S., Mirochnitchenko, O. V., and Pestka, S. (2000). Human cytomegalovirus harbors its own unique IL-10 homolog (cmvIL-10). Proc. Natl Acad. Sci. USA 97, 1695±700. Lai, C. F., Ripperger, J., Morella, K. K., Jurlander, J., Hawley, T. S., Carson, W. E., Kordula, T., Caligiuri, M. A., Hawley, R. G., Fey, G. H., and Baumann, H. (1996). Receptors for interleukin (IL)-10 and IL-6-type cytokines use similar signaling mechanisms for inducing transcription through IL-6 response elements. J. Biol. Chem. 271, 13968±13975. Liu, Y., Wei, S. H., Ho, A. S., de Waal Malefyt, R., and Moore, K. W. (1994). Expression cloning and characterization of a human IL-10 receptor. J. Immunol. 152, 1821±1829. Liu, Y., de Waal Malefyt, R., Briere, F., Parham, C., Bridon, J. M., Banchereau, J., Moore, K. W., and Xu, J. (1997). The EBV IL-10 homologue is a selective agonist with impaired binding to the IL-10 receptor. J. Immunol. 158, 604±613. Michel, G., Mirmohammadsadegh, A., Olasz, E., JarzebskaDeussen, B., Muschen, A., Kemeny, L., Abts, H. F., and Ruzicka, T. (1997). Demonstration and functional analysis of IL-10 receptors in human epidermal cells: decreased expression in psoriatic skin, down-modulation by IL-8, and up-regulation by an antipsoriatic glucocorticosteroid in normal cultured keratinocytes. J. Immunol. 159, 6291±6297. Morikawa, Y., Nishida, H., Misawa, K., Nosaka, T., Miyajima, A., Senba, E., and Kitamura, T. (1998). Induction of synaptosomal-associated protein-23 kD (SNAP-23) by various cytokines. Blood 92, 129±135. O'Farrell, A.-M., Liu, Y., Moore, K. W., and Mui, A. L. (1998). IL-10 inhibits macrophage activation and proliferation by distinct signaling mechanisms: evidence for Stat3-dependent and -independent pathways. EMBO J. 17, 1006±1018.

O'Farrell, A.-M., Parry, D. A., Zindy, F., Roussel, M. F., Lees, E., Moore, K. W., and Mui, A. L.-F. (2000). Stat3-dependent induction of p19INK4D by IL-10 contributes to inhibition of macrophage proliferation. J. Immunol. 164, 4607±4615. Reboul, J., Gardiner, K., Monneron, D., Uze, G., and Lutfalla, G. (1999). Comparative genomic analysis of the interferon/interleukin-10 receptor gene cluster. Genome Res. 9, 242±250. Riley, J. K., Takeda, K., Akira, S., and Schreiber, R. D. (1999). Interleukin-10 receptor signaling through the JAK-STAT pathway. Requirement for two distinct receptor-derived signals for anti-inflammatory action. J. Biol. Chem. 274, 16513±16521. Ripley, B. J., Stephanou, A., Isenberg, D. A., and Latchman, D. S. (1999). Interleukin-10 activates heat-shock protein 90beta gene expression. Immunology 97, 226±231. Spencer, S. D., Di Marco, F., Hooley, J., Pitts-Meek, S., Bauer, M., Ryan, A. M., Sordat, B., Gibbs, V. C., and Aguet, M. (1998). The orphan receptor CRF2±4 is an essential subunit of the interleukin 10 receptor. J. Exp. Med. 187, 571± 578. Song, S., Ling-Hu, H., Roebuck, K. A., Rabbi, M. F., Donnelly, R. P., and Finnegan, A. (1997). Interleukin-10 inhibits interferon-gamma-induced intercellular adhesion molecule-1 gene transcription in human monocytes. Blood 89, 4461±4469. Tan, J. C., Indelicato, S. R., Narula, S. K., Zavodny, P. J., and Chou, C. C. (1993). Characterization of interleukin-10 receptors on human and mouse cells. J. Biol. Chem. 268, 21053±21059. Tan, J. C., Braun, S., Rong, H., DiGiacomo, R., Dolphin, E., Baldwin, S., Narula, S. K., Zavodny, P. J., and Chou, C. C. (1995). Characterization of recombinant extracellular domain of human interleukin-10 receptor. J. Biol. Chem. 270, 12906± 12911. Weber-Nordt, R. M., Meraz, M. A., and Schreiber, R. D. (1994). Lipopolysaccharide-dependent induction of IL-10 receptor expression on murine fibroblasts. J. Immunol. 153, 3734±3744. Weber-Nordt, R. M., Egen, C., Wehinger, J., Ludwig, W., Gouilleux-Gruart, V., Mertelsmann, R., and Finke, J. (1996a). Constitutive activation of STAT proteins in primary lymphoid and myeloid leukemia cells and in Epstein±Barr virus (EBV)related lymphoma cell lines. Blood 88, 809±816. Weber-Nordt, R. M., Riley, J. K., Greenlund, A. C., Moore, K. W., Darnell, J. E., and Schreiber, R. D. (1996b). Stat3 recruitment by two distinct ligand-induced, tyrosinephosphorylated docking sites in the interleukin-10 receptor intracellular domain. J. Biol. Chem. 271, 27954±28961. Wehinger, J., Gouilleux, F., Groner, B., Finke, J., Mertelsmann, R., and Weber-Nordt, R. M. (1996). IL-10 induces DNA binding activity of three STAT proteins (Stat1, Stat3, and Stat5) and their distinct combinatorial assembly in the promoters of selected genes. FEBS Lett. 394, 365±370. Yamaoka, K., Otsuka, T., Niiro, H., Nakashima, H., Tanaka, Y., Nagano, S., Ogami, E., Niho, Y., Hamasaki, N., and Izuhara, K. (1999). Selective DNA-binding activity of interleukin-10-stimulated STAT molecules in human monocytes. J. Interferon Cytokine Res. 19, 679±685. Zdanov, A., Schalk-Hihi, C., and Wlodawer, A. (1996). Crystal structure of human interleukin-10 at 1.6 A resolution and a model of a complex with its soluble receptor. Protein Sci. 5, 1955±1962. Zocchia, C., Spiga, G., Rabin, S. J., Grekova, M., Richert, J., Chernyshev, O., Colton, C., and Mocchetti, I. (1997). Biological activity of interleukin-10 in the central nervous system. Neurochem Int. 30, 433±439.