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. 1996 Feb 1;132(3):299–310. doi: 10.1083/jcb.132.3.299

Role of CD3 gamma in T cell receptor assembly

PMCID: PMC2120718  PMID: 8636209

Abstract

The T cell receptor (TCR) consists of the Ti alpha beta heterodimer and the associated CD3 gamma delta epsilon and zeta 2 chains. The structural relationships between the subunits of the TCR complex are still not fully known. In this study we examined the role of the extracellular (EC), transmembrane (TM), and cytoplasmic (CY) domain of CD3 gamma in assembly and cell surface expression of the complete TCR in human T cells. A computer model indicated that the EC domain of CD3 gamma folds as an Ig domain. Based on this model and on alignment studies, two potential interaction sites were predicted in the EC domain of CD3 gamma. Site-directed mutagenesis demonstrated that these sites play a crucial role in TCR assembly probably by binding to CD3 epsilon. Mutagenesis of N-linked glycosylation sites showed that glycosylation of CD3 gamma is not required for TCR assembly and expression. In contrast, treatment of T cells with tunicamycin suggested that N-linked glycosylation of CD3 delta is required for TCR assembly. Site-directed mutagenesis of the acidic amino acid in the TM domain of CD3 gamma demonstrated that this residue is involved in TCR assembly probably by binding to Ti beta. Deletion of the entire CY domain of CD3 gamma did not prevent assembly and expression of the TCR. In conclusion, this study demonstrated that specific TCR interaction sites exist in both the EC and TM domain of CD3 gamma. Furthermore, the study indicated that, in contrast to CD3 gamma, glycosylation of CD3 delta is required for TCR assembly and expression.

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Selected References

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  1. Alcover A., Mariuzza R. A., Ermonval M., Acuto O. Lysine 271 in the transmembrane domain of the T-cell antigen receptor beta chain is necessary for its assembly with the CD3 complex but not for alpha/beta dimerization. J Biol Chem. 1990 Mar 5;265(7):4131–4135. [PubMed] [Google Scholar]
  2. Berkhout B., Alarcon B., Terhorst C. Transfection of genes encoding the T cell receptor-associated CD3 complex into COS cells results in assembly of the macromolecular structure. J Biol Chem. 1988 Jun 15;263(17):8528–8536. [PubMed] [Google Scholar]
  3. Blumberg R. S., Alarcon B., Sancho J., McDermott F. V., Lopez P., Breitmeyer J., Terhorst C. Assembly and function of the T cell antigen receptor. Requirement of either the lysine or arginine residues in the transmembrane region of the alpha chain. J Biol Chem. 1990 Aug 15;265(23):14036–14043. [PubMed] [Google Scholar]
  4. Bonifacino J. S., Chen C., Lippincott-Schwartz J., Ashwell J. D., Klausner R. D. Subunit interactions within the T-cell antigen receptor: clues from the study of partial complexes. Proc Natl Acad Sci U S A. 1988 Sep;85(18):6929–6933. doi: 10.1073/pnas.85.18.6929. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Bonifacino J. S., Suzuki C. K., Lippincott-Schwartz J., Weissman A. M., Klausner R. D. Pre-Golgi degradation of newly synthesized T-cell antigen receptor chains: intrinsic sensitivity and the role of subunit assembly. J Cell Biol. 1989 Jul;109(1):73–83. doi: 10.1083/jcb.109.1.73. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Brenner M. B., Trowbridge I. S., Strominger J. L. Cross-linking of human T cell receptor proteins: association between the T cell idiotype beta subunit and the T3 glycoprotein heavy subunit. Cell. 1985 Jan;40(1):183–190. doi: 10.1016/0092-8674(85)90321-6. [DOI] [PubMed] [Google Scholar]
  7. Buferne M., Luton F., Letourneur F., Hoeveler A., Couez D., Barad M., Malissen B., Schmitt-Verhulst A. M., Boyer C. Role of CD3 delta in surface expression of the TCR/CD3 complex and in activation for killing analyzed with a CD3 delta-negative cytotoxic T lymphocyte variant. J Immunol. 1992 Feb 1;148(3):657–664. [PubMed] [Google Scholar]
  8. Carson G. R., Kuestner R. E., Ahmed A., Pettey C. L., Concino M. F. Six chains of the human T cell antigen receptor.CD3 complex are necessary and sufficient for processing the receptor heterodimer to the cell surface. J Biol Chem. 1991 Apr 25;266(12):7883–7887. [PubMed] [Google Scholar]
  9. Caspar-Bauguil S., Arnaud J., Gouaillard C., Hou X., Geisler C., Rubin B. Functionally important amino acids in the TCR revealed by immunoselection of membrane TCR-negative T cells. J Immunol. 1994 Jun 1;152(11):5288–5298. [PubMed] [Google Scholar]
  10. Chothia C., Boswell D. R., Lesk A. M. The outline structure of the T-cell alpha beta receptor. EMBO J. 1988 Dec 1;7(12):3745–3755. doi: 10.1002/j.1460-2075.1988.tb03258.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Chou P. Y., Fasman G. D. Prediction of beta-turns. Biophys J. 1979 Jun;26(3):367–383. doi: 10.1016/S0006-3495(79)85259-5. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Clevers H., Alarcon B., Wileman T., Terhorst C. The T cell receptor/CD3 complex: a dynamic protein ensemble. Annu Rev Immunol. 1988;6:629–662. doi: 10.1146/annurev.iy.06.040188.003213. [DOI] [PubMed] [Google Scholar]
  13. Cosson P., Lankford S. P., Bonifacino J. S., Klausner R. D. Membrane protein association by potential intramembrane charge pairs. Nature. 1991 May 30;351(6325):414–416. doi: 10.1038/351414a0. [DOI] [PubMed] [Google Scholar]
  14. Dembić Z., Haas W., Weiss S., McCubrey J., Kiefer H., von Boehmer H., Steinmetz M. Transfer of specificity by murine alpha and beta T-cell receptor genes. Nature. 1986 Mar 20;320(6059):232–238. doi: 10.1038/320232a0. [DOI] [PubMed] [Google Scholar]
  15. Dietrich J., Hou X., Wegener A. M., Geisler C. CD3 gamma contains a phosphoserine-dependent di-leucine motif involved in down-regulation of the T cell receptor. EMBO J. 1994 May 1;13(9):2156–2166. doi: 10.1002/j.1460-2075.1994.tb06492.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Dill K. A. Dominant forces in protein folding. Biochemistry. 1990 Aug 7;29(31):7133–7155. doi: 10.1021/bi00483a001. [DOI] [PubMed] [Google Scholar]
  17. Geisler C. Failure to synthesize the CD3-gamma chain. Consequences for T cell antigen receptor assembly, processing, and expression. J Immunol. 1992 Apr 15;148(8):2437–2445. [PubMed] [Google Scholar]
  18. Geisler C., Plesner T., Pallesen G., Skjødt K., Odum N., Larsen J. K. Characterization and expression of the human T cell receptor-T3 complex by monoclonal antibody F101.01. Scand J Immunol. 1988 Jun;27(6):685–696. doi: 10.1111/j.1365-3083.1988.tb02402.x. [DOI] [PubMed] [Google Scholar]
  19. Geisler C., Rubin B., Caspar-Bauguil S., Champagne E., Vangsted A., Hou X., Gajhede M. Structural mutations of C-domains in members of the Ig superfamily. Consequences for the interactions between the T cell antigen receptor and the zeta 2 homodimer. J Immunol. 1992 Jun 1;148(11):3469–3477. [PubMed] [Google Scholar]
  20. Geisler C., Schøller J., Wahi M. A., Rubin B., Weiss A. Association of the human CD3-zeta chain with the alpha beta-T cell receptor/CD3 complex. Clues from a T cell variant with a mutated T cell receptor-alpha chain. J Immunol. 1990 Sep 15;145(6):1761–1767. [PubMed] [Google Scholar]
  21. Gold D. P., Clevers H., Alarcon B., Dunlap S., Novotny J., Williams A. F., Terhorst C. Evolutionary relationship between the T3 chains of the T-cell receptor complex and the immunoglobulin supergene family. Proc Natl Acad Sci U S A. 1987 Nov;84(21):7649–7653. doi: 10.1073/pnas.84.21.7649. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Hall C., Berkhout B., Alarcon B., Sancho J., Wileman T., Terhorst C. Requirements for cell surface expression of the human TCR/CD3 complex in non-T cells. Int Immunol. 1991 Apr;3(4):359–368. doi: 10.1093/intimm/3.4.359. [DOI] [PubMed] [Google Scholar]
  23. Hou X., Dietrich J., Kuhlmann J., Wegener A. M., Geisler C. Structure of the T cell receptor in a Ti alpha V beta 2, alpha V beta 8-positive T cell line. Eur J Immunol. 1994 May;24(5):1228–1233. doi: 10.1002/eji.1830240534. [DOI] [PubMed] [Google Scholar]
  24. Janin J., Chothia C. The structure of protein-protein recognition sites. J Biol Chem. 1990 Sep 25;265(27):16027–16030. [PubMed] [Google Scholar]
  25. John S., Banting G. S., Goodfellow P. N., Owen M. J. Surface expression of the T cell receptor complex requires charged residues within the alpha chain transmembrane region. Eur J Immunol. 1989 Feb;19(2):335–339. doi: 10.1002/eji.1830190218. [DOI] [PubMed] [Google Scholar]
  26. Kabsch W., Sander C. Dictionary of protein secondary structure: pattern recognition of hydrogen-bonded and geometrical features. Biopolymers. 1983 Dec;22(12):2577–2637. doi: 10.1002/bip.360221211. [DOI] [PubMed] [Google Scholar]
  27. Kearse K. P., Williams D. B., Singer A. Persistence of glucose residues on core oligosaccharides prevents association of TCR alpha and TCR beta proteins with calnexin and results specifically in accelerated degradation of nascent TCR alpha proteins within the endoplasmic reticulum. EMBO J. 1994 Aug 15;13(16):3678–3686. doi: 10.1002/j.1460-2075.1994.tb06677.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Kim K. M., Adachi T., Nielsen P. J., Terashima M., Lamers M. C., Köhler G., Reth M. Two new proteins preferentially associated with membrane immunoglobulin D. EMBO J. 1994 Aug 15;13(16):3793–3800. doi: 10.1002/j.1460-2075.1994.tb06690.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Koning F., Maloy W. L., Coligan J. E. The implications of subunit interactions for the structure of the T cell receptor-CD3 complex. Eur J Immunol. 1990 Feb;20(2):299–305. doi: 10.1002/eji.1830200211. [DOI] [PubMed] [Google Scholar]
  30. Kornfeld R., Kornfeld S. Assembly of asparagine-linked oligosaccharides. Annu Rev Biochem. 1985;54:631–664. doi: 10.1146/annurev.bi.54.070185.003215. [DOI] [PubMed] [Google Scholar]
  31. Krissansen G. W., Owen M. J., Verbi W., Crumpton M. J. Primary structure of the T3 gamma subunit of the T3/T cell antigen receptor complex deduced from cDNA sequences: evolution of the T3 gamma and delta subunits. EMBO J. 1986 Aug;5(8):1799–1808. doi: 10.1002/j.1460-2075.1986.tb04429.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Kuhlmann J., Geisler C. Structure of the T-cell receptor in a Ti alpha beta, Ti gamma delta double positive T-cell line. Scand J Immunol. 1993 Feb;37(2):271–275. doi: 10.1111/j.1365-3083.1993.tb01766.x. [DOI] [PubMed] [Google Scholar]
  33. Kyte J., Doolittle R. F. A simple method for displaying the hydropathic character of a protein. J Mol Biol. 1982 May 5;157(1):105–132. doi: 10.1016/0022-2836(82)90515-0. [DOI] [PubMed] [Google Scholar]
  34. Levins B., Colaco C. A. The CD3 gamma-chain, transmembrane signalling or recognition? Immunogenetics. 1992;35(2):140–143. doi: 10.1007/BF00189525. [DOI] [PubMed] [Google Scholar]
  35. Malissen B., Schmitt-Verhulst A. M. Transmembrane signalling through the T-cell-receptor-CD3 complex. Curr Opin Immunol. 1993 Jun;5(3):324–333. doi: 10.1016/0952-7915(93)90049-x. [DOI] [PubMed] [Google Scholar]
  36. Manolios N., Bonifacino J. S., Klausner R. D. Transmembrane helical interactions and the assembly of the T cell receptor complex. Science. 1990 Jul 20;249(4966):274–277. doi: 10.1126/science.2142801. [DOI] [PubMed] [Google Scholar]
  37. Manolios N., Kemp O., Li Z. G. The T cell antigen receptor alpha and beta chains interact via distinct regions with CD3 chains. Eur J Immunol. 1994 Jan;24(1):84–92. doi: 10.1002/eji.1830240114. [DOI] [PubMed] [Google Scholar]
  38. Manolios N., Letourneur F., Bonifacino J. S., Klausner R. D. Pairwise, cooperative and inhibitory interactions describe the assembly and probable structure of the T-cell antigen receptor. EMBO J. 1991 Jul;10(7):1643–1651. doi: 10.1002/j.1460-2075.1991.tb07687.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Marquart M., Deisenhofer J., Huber R., Palm W. Crystallographic refinement and atomic models of the intact immunoglobulin molecule Kol and its antigen-binding fragment at 3.0 A and 1.0 A resolution. J Mol Biol. 1980 Aug 25;141(4):369–391. doi: 10.1016/0022-2836(80)90252-1. [DOI] [PubMed] [Google Scholar]
  40. Padovan E., Casorati G., Dellabona P., Meyer S., Brockhaus M., Lanzavecchia A. Expression of two T cell receptor alpha chains: dual receptor T cells. Science. 1993 Oct 15;262(5132):422–424. doi: 10.1126/science.8211163. [DOI] [PubMed] [Google Scholar]
  41. Punt J. A., Roberts J. L., Kearse K. P., Singer A. Stoichiometry of the T cell antigen receptor (TCR) complex: each TCR/CD3 complex contains one TCR alpha, one TCR beta, and two CD3 epsilon chains. J Exp Med. 1994 Aug 1;180(2):587–593. doi: 10.1084/jem.180.2.587. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Ryu S. E., Kwong P. D., Truneh A., Porter T. G., Arthos J., Rosenberg M., Dai X. P., Xuong N. H., Axel R., Sweet R. W. Crystal structure of an HIV-binding recombinant fragment of human CD4. Nature. 1990 Nov 29;348(6300):419–426. doi: 10.1038/348419a0. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Saito T., Weiss A., Miller J., Norcross M. A., Germain R. N. Specific antigen-Ia activation of transfected human T cells expressing murine Ti alpha beta-human T3 receptor complexes. Nature. 1987 Jan 8;325(7000):125–130. doi: 10.1038/325125a0. [DOI] [PubMed] [Google Scholar]
  44. Singer S. J. The structure and insertion of integral proteins in membranes. Annu Rev Cell Biol. 1990;6:247–296. doi: 10.1146/annurev.cb.06.110190.001335. [DOI] [PubMed] [Google Scholar]
  45. Transy C., Moingeon P., Stebbins C., Reinherz E. L. Deletion of the cytoplasmic region of the CD3 epsilon subunit does not prevent assembly of a functional T-cell receptor. Proc Natl Acad Sci U S A. 1989 Sep;86(18):7108–7112. doi: 10.1073/pnas.86.18.7108. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. Van Neerven J., Coligan J. E., Koning F. Structural comparison of alpha/beta and gamma/delta T cell receptor-CD3 complexes reveals identical subunit interactions but distinct cross-linking patterns of T cell receptor chains. Eur J Immunol. 1990 Sep;20(9):2105–2111. doi: 10.1002/eji.1830200932. [DOI] [PubMed] [Google Scholar]
  47. Varki A. Biological roles of oligosaccharides: all of the theories are correct. Glycobiology. 1993 Apr;3(2):97–130. doi: 10.1093/glycob/3.2.97. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Venkatachalam C. M., Ramachandran G. N. Conformation of polypeptide chains. Annu Rev Biochem. 1969;38:45–82. doi: 10.1146/annurev.bi.38.070169.000401. [DOI] [PubMed] [Google Scholar]
  49. Wang J. H., Yan Y. W., Garrett T. P., Liu J. H., Rodgers D. W., Garlick R. L., Tarr G. E., Husain Y., Reinherz E. L., Harrison S. C. Atomic structure of a fragment of human CD4 containing two immunoglobulin-like domains. Nature. 1990 Nov 29;348(6300):411–418. doi: 10.1038/348411a0. [DOI] [PubMed] [Google Scholar]
  50. Wange R. L., Malek S. N., Desiderio S., Samelson L. E. Tandem SH2 domains of ZAP-70 bind to T cell antigen receptor zeta and CD3 epsilon from activated Jurkat T cells. J Biol Chem. 1993 Sep 15;268(26):19797–19801. [PubMed] [Google Scholar]
  51. Wegener A. M., Hou X., Dietrich J., Geisler C. Distinct domains of the CD3-gamma chain are involved in surface expression and function of the T cell antigen receptor. J Biol Chem. 1995 Mar 3;270(9):4675–4680. doi: 10.1074/jbc.270.9.4675. [DOI] [PubMed] [Google Scholar]
  52. Wegener A. M., Letourneur F., Hoeveler A., Brocker T., Luton F., Malissen B. The T cell receptor/CD3 complex is composed of at least two autonomous transduction modules. Cell. 1992 Jan 10;68(1):83–95. doi: 10.1016/0092-8674(92)90208-t. [DOI] [PubMed] [Google Scholar]
  53. Wileman T., Kane L. P., Young J., Carson G. R., Terhorst C. Associations between subunit ectodomains promote T cell antigen receptor assembly and protect against degradation in the ER. J Cell Biol. 1993 Jul;122(1):67–78. doi: 10.1083/jcb.122.1.67. [DOI] [PMC free article] [PubMed] [Google Scholar]
  54. Williams A. F., Barclay A. N. The immunoglobulin superfamily--domains for cell surface recognition. Annu Rev Immunol. 1988;6:381–405. doi: 10.1146/annurev.iy.06.040188.002121. [DOI] [PubMed] [Google Scholar]
  55. Williams A. M., Enns C. A. A region of the C-terminal portion of the human transferrin receptor contains an asparagine-linked glycosylation site critical for receptor structure and function. J Biol Chem. 1993 Jun 15;268(17):12780–12786. [PubMed] [Google Scholar]

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