Skip to main content
The Journal of Experimental Medicine logoLink to The Journal of Experimental Medicine
. 1989 Jan 1;169(1):115–133. doi: 10.1084/jem.169.1.115

Molecular genetic analysis of 178 I-Abm12-reactive T cells

PMCID: PMC2189175  PMID: 2783331

Abstract

We have studied the genetic diversity of the TCR repertoire to the murine alloantigen I-Abm12 by generating a panel of 178 C57BL/10- derived I-Abm12-reactive T cell hybridomas. The expression of V alpha and V beta gene families was examined in this panel and the frequency of expression of V beta, but not ofV alpha, gene families differed significantly from that observed in a companion panel of random C57BL/10-derived hybridomas. The V beta 5 gene family was expressed significantly less frequently while the V beta 14, V beta 15, and V beta 16 genes were expressed significantly more frequently in the panel of I-Abm12-reactive than in the panel of random hybridomas. The junctional regions (VJ alpha and VDJ beta) of TCR V alpha and V beta genes from selected I-Abm12-specific hybridomas were amplified using the polymerase chain reaction, and directly sequenced. Surprisingly, no conserved J alpha, D beta, J beta, or N region-encoded sequences among these selected I-Abm12-reactive TCRs were identified. Thus, the T cell response to an I-A alloantigen that differs by only three amino acid residues from the I-A molecule of the responding strain is genetically complex but nonrandom. We have estimated that the repertoire to this alloantigen is comprised of at least 37 different TCRs.

Full Text

The Full Text of this article is available as a PDF (1.3 MB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Arden B., Klotz J. L., Siu G., Hood L. E. Diversity and structure of genes of the alpha family of mouse T-cell antigen receptor. 1985 Aug 29-Sep 4Nature. 316(6031):783–787. doi: 10.1038/316783a0. [DOI] [PubMed] [Google Scholar]
  2. Ashwell J. D., Chen C., Schwartz R. H. High frequency and nonrandom distribution of alloreactivity in T cell clones selected for recognition of foreign antigen in association with self class II molecules. J Immunol. 1986 Jan;136(2):389–395. [PubMed] [Google Scholar]
  3. Babbitt B. P., Allen P. M., Matsueda G., Haber E., Unanue E. R. Binding of immunogenic peptides to Ia histocompatibility molecules. 1985 Sep 26-Oct 2Nature. 317(6035):359–361. doi: 10.1038/317359a0. [DOI] [PubMed] [Google Scholar]
  4. Barth R. K., Kim B. S., Lan N. C., Hunkapiller T., Sobieck N., Winoto A., Gershenfeld H., Okada C., Hansburg D., Weissman I. L. The murine T-cell receptor uses a limited repertoire of expressed V beta gene segments. Nature. 1985 Aug 8;316(6028):517–523. doi: 10.1038/316517a0. [DOI] [PubMed] [Google Scholar]
  5. Behlke M. A., Chou H. S., Huppi K., Loh D. Y. Murine T-cell receptor mutants with deletions of beta-chain variable region genes. Proc Natl Acad Sci U S A. 1986 Feb;83(3):767–771. doi: 10.1073/pnas.83.3.767. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Bevan M. J., Fink P. J. The influence of thymus H-2 antigens on the specificity of maturing killer and helper cells. Immunol Rev. 1978;42:3–19. doi: 10.1111/j.1600-065x.1978.tb00256.x. [DOI] [PubMed] [Google Scholar]
  7. Bjorkman P. J., Saper M. A., Samraoui B., Bennett W. S., Strominger J. L., Wiley D. C. Structure of the human class I histocompatibility antigen, HLA-A2. Nature. 1987 Oct 8;329(6139):506–512. doi: 10.1038/329506a0. [DOI] [PubMed] [Google Scholar]
  8. Bjorkman P. J., Saper M. A., Samraoui B., Bennett W. S., Strominger J. L., Wiley D. C. The foreign antigen binding site and T cell recognition regions of class I histocompatibility antigens. Nature. 1987 Oct 8;329(6139):512–518. doi: 10.1038/329512a0. [DOI] [PubMed] [Google Scholar]
  9. Blackman M. A., Kappler J. W., Marrack P. T-cell specificity and repertoire. Immunol Rev. 1988 Jan;101:5–19. doi: 10.1111/j.1600-065x.1988.tb00730.x. [DOI] [PubMed] [Google Scholar]
  10. Born W., Yagüe J., Palmer E., Kappler J., Marrack P. Rearrangement of T-cell receptor beta-chain genes during T-cell development. Proc Natl Acad Sci U S A. 1985 May;82(9):2925–2929. doi: 10.1073/pnas.82.9.2925. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Brown J. H., Jardetzky T., Saper M. A., Samraoui B., Bjorkman P. J., Wiley D. C. A hypothetical model of the foreign antigen binding site of class II histocompatibility molecules. Nature. 1988 Apr 28;332(6167):845–850. doi: 10.1038/332845a0. [DOI] [PubMed] [Google Scholar]
  12. Buus S., Sette A., Colon S. M., Jenis D. M., Grey H. M. Isolation and characterization of antigen-Ia complexes involved in T cell recognition. Cell. 1986 Dec 26;47(6):1071–1077. doi: 10.1016/0092-8674(86)90822-6. [DOI] [PubMed] [Google Scholar]
  13. Cheley S., Anderson R. A reproducible microanalytical method for the detection of specific RNA sequences by dot-blot hybridization. Anal Biochem. 1984 Feb;137(1):15–19. doi: 10.1016/0003-2697(84)90339-7. [DOI] [PubMed] [Google Scholar]
  14. Cohn M. The T-cell receptor mediating restrictive recognition of antigen. Cell. 1983 Jul;33(3):657–669. doi: 10.1016/0092-8674(83)90009-0. [DOI] [PubMed] [Google Scholar]
  15. Feinberg A. P., Vogelstein B. A technique for radiolabeling DNA restriction endonuclease fragments to high specific activity. Anal Biochem. 1983 Jul 1;132(1):6–13. doi: 10.1016/0003-2697(83)90418-9. [DOI] [PubMed] [Google Scholar]
  16. Fink P. J., Matis L. A., McElligott D. L., Bookman M., Hedrick S. M. Correlations between T-cell specificity and the structure of the antigen receptor. Nature. 1986 May 15;321(6067):219–226. doi: 10.1038/321219a0. [DOI] [PubMed] [Google Scholar]
  17. Garman R. D., Ko J. L., Vulpe C. D., Raulet D. H. T-cell receptor variable region gene usage in T-cell populations. Proc Natl Acad Sci U S A. 1986 Jun;83(11):3987–3991. doi: 10.1073/pnas.83.11.3987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Goverman J., Minard K., Shastri N., Hunkapiller T., Hansburg D., Sercarz E., Hood L. Rearranged beta T cell receptor genes in a helper T cell clone specific for lysozyme: no correlation between V beta and MHC restriction. Cell. 1985 Apr;40(4):859–867. doi: 10.1016/0092-8674(85)90345-9. [DOI] [PubMed] [Google Scholar]
  19. Guillet J. G., Lai M. Z., Briner T. J., Buus S., Sette A., Grey H. M., Smith J. A., Gefter M. L. Immunological self, nonself discrimination. Science. 1987 Feb 20;235(4791):865–870. doi: 10.1126/science.2433769. [DOI] [PubMed] [Google Scholar]
  20. Hünig T. R., Bevan M. J. Antigen recognition by cloned cytotoxic T lymphocytes follows rules predicted by the altered-self hypothesis. J Exp Med. 1982 Jan 1;155(1):111–125. doi: 10.1084/jem.155.1.111. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Iwamoto A., Ohashi P. S., Pircher H., Walker C. L., Michalopoulos E. E., Rupp F., Hengartner H., Mak T. W. T cell receptor variable gene usage in a specific cytotoxic T cell response. Primary structure of the antigen-MHC receptor of four hapten-specific cytotoxic T cell clones. J Exp Med. 1987 Mar 1;165(3):591–600. doi: 10.1084/jem.165.3.591. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Kappler J. W., Roehm N., Marrack P. T cell tolerance by clonal elimination in the thymus. Cell. 1987 Apr 24;49(2):273–280. doi: 10.1016/0092-8674(87)90568-x. [DOI] [PubMed] [Google Scholar]
  23. Kappler J. W., Skidmore B., White J., Marrack P. Antigen-inducible, H-2-restricted, interleukin-2-producing T cell hybridomas. Lack of independent antigen and H-2 recognition. J Exp Med. 1981 May 1;153(5):1198–1214. doi: 10.1084/jem.153.5.1198. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Kappler J. W., Wade T., White J., Kushnir E., Blackman M., Bill J., Roehm N., Marrack P. A T cell receptor V beta segment that imparts reactivity to a class II major histocompatibility complex product. Cell. 1987 Apr 24;49(2):263–271. doi: 10.1016/0092-8674(87)90567-8. [DOI] [PubMed] [Google Scholar]
  25. Kappler J., White J., Wegmann D., Mustain E., Marrack P. Antigen presentation by Ia+ B cell hybridomas to H-2-restricted T cell hybridomas. Proc Natl Acad Sci U S A. 1982 Jun;79(11):3604–3607. doi: 10.1073/pnas.79.11.3604. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Klein J., Figueroa F. Polymorphism of the mouse H-2 loci. Immunol Rev. 1981;60:23–57. doi: 10.1111/j.1600-065x.1981.tb00361.x. [DOI] [PubMed] [Google Scholar]
  27. Lafuse W. P., McCormick J. F., Melvold R. W., David C. S. Serological and biochemical analysis of Ia molecules in the I-A mutant B6.C-H-2. Transplantation. 1981 Jun;31(6):434–438. doi: 10.1097/00007890-198106000-00006. [DOI] [PubMed] [Google Scholar]
  28. Lin C. C., Rosenthal A. S., Passmore H. C., Hansen T. H. Selective loss of antigen-specific Ir gene function in IA mutant B6.C-H-2bm12 is an antigen presenting cell defect. Proc Natl Acad Sci U S A. 1981 Oct;78(10):6406–6410. doi: 10.1073/pnas.78.10.6406. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Lindahl K. F., Wilson D. B. Histocompatibility antigen-activated cytotoxic T lymphocytes. II. Estimates of the frequency and specificity of precursors. J Exp Med. 1977 Mar 1;145(3):508–522. doi: 10.1084/jem.145.3.508. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Marrack P., Kappler J. T cells can distinguish between allogeneic major histocompatibility complex products on different cell types. Nature. 1988 Apr 28;332(6167):840–843. doi: 10.1038/332840a0. [DOI] [PubMed] [Google Scholar]
  31. McElligott D. L., Sorger S. B., Matis L. A., Hedrick S. M. Two distinct mechanisms account for the immune response (Ir) gene control of the T cell response to pigeon cytochrome c. J Immunol. 1988 Jun 15;140(12):4123–4131. [PubMed] [Google Scholar]
  32. McKenzie I. F., Morgan G. M., Sandrin M. S., Michaelides M. M., Melvold R. W., Kohn H. I. B6.C-H-2bm12. A new H-2 mutation in the I region in the mouse. J Exp Med. 1979 Dec 1;150(6):1323–1338. doi: 10.1084/jem.150.6.1323. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Mengle-Gaw L., Conner S., McDevitt H. O., Fathman C. G. Gene conversion between murine class II major histocompatibility complex loci. Functional and molecular evidence from the bm 12 mutant. J Exp Med. 1984 Oct 1;160(4):1184–1194. doi: 10.1084/jem.160.4.1184. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Michaelides M., Sandrin M., Morgan G., McKenzie I. F., Ashman R., Melvold R. W. Ir gene function in an I-A subregion mutant B6.C-H-2bm12. J Exp Med. 1981 Feb 1;153(2):464–469. doi: 10.1084/jem.153.2.464. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Mullis K. B., Faloona F. A. Specific synthesis of DNA in vitro via a polymerase-catalyzed chain reaction. Methods Enzymol. 1987;155:335–350. doi: 10.1016/0076-6879(87)55023-6. [DOI] [PubMed] [Google Scholar]
  36. Reimann J., Bellan A. Use of V beta.8 genes in splenic Lyt-2+ cytotoxic lymphocyte precursors reactive to bm1 or bm14 alloantigen in individual C57BL/6 mice. Eur J Immunol. 1986 Dec;16(12):1597–1602. doi: 10.1002/eji.1830161220. [DOI] [PubMed] [Google Scholar]
  37. Rigby P. W., Dieckmann M., Rhodes C., Berg P. Labeling deoxyribonucleic acid to high specific activity in vitro by nick translation with DNA polymerase I. J Mol Biol. 1977 Jun 15;113(1):237–251. doi: 10.1016/0022-2836(77)90052-3. [DOI] [PubMed] [Google Scholar]
  38. Saiki R. K., Gelfand D. H., Stoffel S., Scharf S. J., Higuchi R., Horn G. T., Mullis K. B., Erlich H. A. Primer-directed enzymatic amplification of DNA with a thermostable DNA polymerase. Science. 1988 Jan 29;239(4839):487–491. doi: 10.1126/science.2448875. [DOI] [PubMed] [Google Scholar]
  39. Saiki R. K., Scharf S., Faloona F., Mullis K. B., Horn G. T., Erlich H. A., Arnheim N. Enzymatic amplification of beta-globin genomic sequences and restriction site analysis for diagnosis of sickle cell anemia. Science. 1985 Dec 20;230(4732):1350–1354. doi: 10.1126/science.2999980. [DOI] [PubMed] [Google Scholar]
  40. Sanger F., Nicklen S., Coulson A. R. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. doi: 10.1073/pnas.74.12.5463. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Sherman D. H., Hochman P. S., Dick R., Tizard R., Ramachandran K. L., Flavell R. A., Huber B. T. Molecular analysis of antigen recognition by insulin-specific T-cell hybridomas from B6 wild-type and bm12 mutant mice. Mol Cell Biol. 1987 May;7(5):1865–1872. doi: 10.1128/mcb.7.5.1865. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Sherman L. A. Dissection of the B10.D2 anti-H-2Kb cytolytic T lymphocyte receptor repertoire. J Exp Med. 1980 Jun 1;151(6):1386–1397. doi: 10.1084/jem.151.6.1386. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Sherman L. A. Influence of the major histocompatibility complex on the repertoire of allospecific cytolytic T lymphocytes. J Exp Med. 1982 Feb 1;155(2):380–389. doi: 10.1084/jem.155.2.380. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Teh H. S., Harley E., Phillips R. A., Miller R. G. Quantitative studies on the precursors of cytotoxic lymphocytes. I. Characterization of a clonal assay and determination of the size of clones derived from single precursors. J Immunol. 1977 Mar;118(3):1049–1056. [PubMed] [Google Scholar]
  45. Tse H. Y., Kanamori S., Walsh W. D., Hansen T. H. The murine bm12 gene conversion provides evidence that T cells recognize predominantly Ia conformation. Proc Natl Acad Sci U S A. 1985 Oct;82(20):7058–7062. doi: 10.1073/pnas.82.20.7058. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. Wilson R. K., Lai E., Concannon P., Barth R. K., Hood L. E. Structure, organization and polymorphism of murine and human T-cell receptor alpha and beta chain gene families. Immunol Rev. 1988 Jan;101:149–172. doi: 10.1111/j.1600-065x.1988.tb00736.x. [DOI] [PubMed] [Google Scholar]
  47. Zinkernagel R. M., Doherty P. C. Immunological surveillance against altered self components by sensitised T lymphocytes in lymphocytic choriomeningitis. Nature. 1974 Oct 11;251(5475):547–548. doi: 10.1038/251547a0. [DOI] [PubMed] [Google Scholar]

Articles from The Journal of Experimental Medicine are provided here courtesy of The Rockefeller University Press

RESOURCES