Skip to main content
PMC home page
Molecular and Cellular Biology logoLink to Molecular and Cellular Biology
. 1988 Dec;8(12):5459–5469. doi: 10.1128/mcb.8.12.5459

Expression of T-cell receptor alpha-chain genes in transgenic mice.

L J Berg 1, B Fazekas de St Groth 1, F Ivars 1, C C Goodnow 1, S Gilfillan 1, H J Garchon 1, M M Davis 1
PMCID: PMC365649  PMID: 3266655

Abstract

To examine the influences responsible for shaping the T-cell repertoire in vivo, we have introduced T-cell receptors of defined specificity into mice. In this report, we analyze transgenic mice carrying a T-cell receptor alpha-chain gene from a pigeon cytochrome c-reactive T-cell line. A variant of this construct, which has the immunoglobulin heavy-chain enhancer inserted into the JC intron, was also introduced into mice. Addition of the enhancer increased the steady-state level of transgene-encoded mRNA three- to fivefold in cultured T cells, leading to a two- to threefold increase in surface expression. In vivo, the difference between these two constructs was even more significant, increasing the number of transgene-positive cells from approximately 5 to 70% and the T-cell receptor surface density two- to threefold. Surprisingly, while surface expression of either type of transgene was limited to T cells, we found little tissue specificity with respect to transcription. In T cells expressing the alpha chain from the enhancer-containing construct, immunoprecipitation with a 2B4 alpha-specific monoclonal antibody revealed the expected disulfide-linked dimer. Costaining of these T cells with the 2B4 alpha-specific monoclonal antibody versus anti-CD3 indicated that expression of the transgene-encoded alpha chain precludes expression of endogenous alpha chains on the majority of cells; in contrast, 2B4 alpha-chain expression from the construct lacking the enhancer is inefficient at suppressing endogenous alpha-chain expression. In mice of the enhancer lineage, Southern blot analysis indicated suppression of endogenous alpha-chain rearrangements in T-cell populations, consistent with the observed allelic exclusion at the cellular level. Interestingly, newborn, but not adult, mice of this lineage also showed an increase in retention of unrearranged delta-chain loci in thymocyte DNA, presumably resulting from the suppression of alpha-chain rearrangements. This observation indicates that at least a fraction of alpha:beta-positive T cells have never attempted to produce functional delta rearrangements, thus suggesting that alpha:beta and gamma:delta T cells may be derived from different T-cell compartments (at least during the early phases of T-cell differentiation).

Full text

PDF
5466

Images in this article

5461Image
on p.5461
5462Image
on p.5462
5464Image
on p.5464
5466Image
on p.5466

Selected References

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

  1. Banerji J., Olson L., Schaffner W. A lymphocyte-specific cellular enhancer is located downstream of the joining region in immunoglobulin heavy chain genes. Cell. 1983 Jul;33(3):729–740. doi: 10.1016/0092-8674(83)90015-6. [DOI] [PubMed] [Google Scholar]
  2. Becker D. M., Pattern P., Chien Y., Yokota T., Eshhar Z., Giedlin M., Gascoigne N. R., Goodnow C., Wolf R., Arai K. Variability and repertoire size of T-cell receptor V alpha gene segments. Nature. 1985 Oct 3;317(6036):430–434. doi: 10.1038/317430a0. [DOI] [PubMed] [Google Scholar]
  3. Brawerman G., Mendecki J., Lee S. Y. A procedure for the isolation of mammalian messenger ribonucleic acid. Biochemistry. 1972 Feb 15;11(4):637–641. doi: 10.1021/bi00754a027. [DOI] [PubMed] [Google Scholar]
  4. Brinster R. L., Chen H. Y., Trumbauer M., Senear A. W., Warren R., Palmiter R. D. Somatic expression of herpes thymidine kinase in mice following injection of a fusion gene into eggs. Cell. 1981 Nov;27(1 Pt 2):223–231. doi: 10.1016/0092-8674(81)90376-7. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Buus S., Sette A., Colon S. M., Miles C., Grey H. M. The relation between major histocompatibility complex (MHC) restriction and the capacity of Ia to bind immunogenic peptides. Science. 1987 Mar 13;235(4794):1353–1358. doi: 10.1126/science.2435001. [DOI] [PubMed] [Google Scholar]
  6. Chen S., Botteri F., van der Putten H., Landel C. P., Evans G. A. A lymphoproliferative abnormality associated with inappropriate expression of the Thy-1 antigen in transgenic mice. Cell. 1987 Oct 9;51(1):7–19. doi: 10.1016/0092-8674(87)90005-5. [DOI] [PubMed] [Google Scholar]
  7. Chien Y. H., Iwashima M., Kaplan K. B., Elliott J. F., Davis M. M. A new T-cell receptor gene located within the alpha locus and expressed early in T-cell differentiation. 1987 Jun 25-Jul 1Nature. 327(6124):677–682. doi: 10.1038/327677a0. [DOI] [PubMed] [Google Scholar]
  8. Chirgwin J. M., Przybyla A. E., MacDonald R. J., Rutter W. J. Isolation of biologically active ribonucleic acid from sources enriched in ribonuclease. Biochemistry. 1979 Nov 27;18(24):5294–5299. doi: 10.1021/bi00591a005. [DOI] [PubMed] [Google Scholar]
  9. Chou H. S., Behlke M. A., Godambe S. A., Russell J. H., Brooks C. G., Loh D. Y. T cell receptor genes in an alloreactive CTL clone: implications for rearrangement and germline diversity of variable gene segments. EMBO J. 1986 Sep;5(9):2149–2155. doi: 10.1002/j.1460-2075.1986.tb04478.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Coffman R. L., Weissman I. L. B220: a B cell-specific member of th T200 glycoprotein family. Nature. 1981 Feb 19;289(5799):681–683. doi: 10.1038/289681a0. [DOI] [PubMed] [Google Scholar]
  11. Costantini F., Lacy E. Introduction of a rabbit beta-globin gene into the mouse germ line. Nature. 1981 Nov 5;294(5836):92–94. doi: 10.1038/294092a0. [DOI] [PubMed] [Google Scholar]
  12. Deans R. J., Denis K. A., Taylor A., Wall R. Expression of an immunoglobulin heavy chain gene transfected into lymphocytes. Proc Natl Acad Sci U S A. 1984 Mar;81(5):1292–1296. doi: 10.1073/pnas.81.5.1292. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. 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]
  14. Dialynas D. P., Quan Z. S., Wall K. A., Pierres A., Quintáns J., Loken M. R., Pierres M., Fitch F. W. Characterization of the murine T cell surface molecule, designated L3T4, identified by monoclonal antibody GK1.5: similarity of L3T4 to the human Leu-3/T4 molecule. J Immunol. 1983 Nov;131(5):2445–2451. [PubMed] [Google Scholar]
  15. Elliott J. F., Rock E. P., Patten P. A., Davis M. M., Chien Y. H. The adult T-cell receptor delta-chain is diverse and distinct from that of fetal thymocytes. Nature. 1988 Feb 18;331(6157):627–631. doi: 10.1038/331627a0. [DOI] [PubMed] [Google Scholar]
  16. Gascoigne N. R., Goodnow C. C., Dudzik K. I., Oi V. T., Davis M. M. Secretion of a chimeric T-cell receptor-immunoglobulin protein. Proc Natl Acad Sci U S A. 1987 May;84(9):2936–2940. doi: 10.1073/pnas.84.9.2936. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Gillies S. D., Morrison S. L., Oi V. T., Tonegawa S. A tissue-specific transcription enhancer element is located in the major intron of a rearranged immunoglobulin heavy chain gene. Cell. 1983 Jul;33(3):717–728. doi: 10.1016/0092-8674(83)90014-4. [DOI] [PubMed] [Google Scholar]
  18. Grosschedl R., Weaver D., Baltimore D., Costantini F. Introduction of a mu immunoglobulin gene into the mouse germ line: specific expression in lymphoid cells and synthesis of functional antibody. Cell. 1984 Oct;38(3):647–658. doi: 10.1016/0092-8674(84)90259-9. [DOI] [PubMed] [Google Scholar]
  19. Haars R., Kronenberg M., Gallatin W. M., Weissman I. L., Owen F. L., Hood L. Rearrangement and expression of T cell antigen receptor and gamma genes during thymic development. J Exp Med. 1986 Jul 1;164(1):1–24. doi: 10.1084/jem.164.1.1. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Hayakawa K., Hardy R. R., Parks D. R., Herzenberg L. A. The "Ly-1 B" cell subpopulation in normal immunodefective, and autoimmune mice. J Exp Med. 1983 Jan 1;157(1):202–218. doi: 10.1084/jem.157.1.202. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Hayday A. C., Diamond D. J., Tanigawa G., Heilig J. S., Folsom V., Saito H., Tonegawa S. Unusual organization and diversity of T-cell receptor alpha-chain genes. 1985 Aug 29-Sep 4Nature. 316(6031):828–832. doi: 10.1038/316828a0. [DOI] [PubMed] [Google Scholar]
  22. Ikuta K., Ogura T., Shimizu A., Honjo T. Low frequency of somatic mutation in beta-chain variable region genes of human T-cell receptors. Proc Natl Acad Sci U S A. 1985 Nov;82(22):7701–7705. doi: 10.1073/pnas.82.22.7701. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Jotereau F., Heuze F., Salomon-Vie V., Gascan H. Cell kinetics in the fetal mouse thymus: precursor cell input, proliferation, and emigration. J Immunol. 1987 Feb 15;138(4):1026–1030. [PubMed] [Google Scholar]
  24. Kemp D. J., Wilson A., Harris A. W., Shortman K. The immunoglobulin mu constant region gene is expressed in mouse thymocytes. Nature. 1980 Jul 10;286(5769):168–170. doi: 10.1038/286168a0. [DOI] [PubMed] [Google Scholar]
  25. Keski-Oja J., Mosher D. F., Vaheri A. Dimeric character of fibronectin, a major cell surface-associated glycoprotein. Biochem Biophys Res Commun. 1977 Jan 24;74(2):699–706. doi: 10.1016/0006-291x(77)90359-x. [DOI] [PubMed] [Google Scholar]
  26. Krimpenfort P., de Jong R., Uematsu Y., Dembic Z., Ryser S., von Boehmer H., Steinmetz M., Berns A. Transcription of T cell receptor beta-chain genes is controlled by a downstream regulatory element. EMBO J. 1988 Mar;7(3):745–750. doi: 10.1002/j.1460-2075.1988.tb02871.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Ledbetter J. A., Herzenberg L. A. Xenogeneic monoclonal antibodies to mouse lymphoid differentiation antigens. Immunol Rev. 1979;47:63–90. doi: 10.1111/j.1600-065x.1979.tb00289.x. [DOI] [PubMed] [Google Scholar]
  28. Lee N. E., Davis M. M. T cell receptor beta-chain genes in BW5147 and other AKR tumors. Deletion order of murine V beta gene segments and possible 5' regulatory regions. J Immunol. 1988 Mar 1;140(5):1665–1675. [PubMed] [Google Scholar]
  29. Leiden J. M., Dialynas D. P., Duby A. D., Murre C., Seidman J., Strominger J. L. Rearrangement and expression of T-cell antigen receptor genes in human T-lymphocyte tumor lines and normal human T-cell clones: evidence for allelic exclusion of Ti beta gene expression and preferential use of a J beta 2 gene segment. Mol Cell Biol. 1986 Sep;6(9):3207–3214. doi: 10.1128/mcb.6.9.3207. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Lindsten T., Fowlkes B. J., Samelson L. E., Davis M. M., Chien Y. H. Transient rearrangements of the T cell antigen receptor alpha locus in early thymocytes. J Exp Med. 1987 Sep 1;166(3):761–775. doi: 10.1084/jem.166.3.761. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Loken M. R., Stall A. M. Flow cytometry as an analytical and preparative tool in immunology. J Immunol Methods. 1982;50(3):R85–112. doi: 10.1016/0022-1759(82)90161-2. [DOI] [PubMed] [Google Scholar]
  32. Luria S., Gross G., Horowitz M., Givol D. Promoter and enhancer elements in the rearranged alpha chain gene of the human T cell receptor. EMBO J. 1987 Nov;6(11):3307–3312. doi: 10.1002/j.1460-2075.1987.tb02650.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Mason J. O., Williams G. T., Neuberger M. S. Transcription cell type specificity is conferred by an immunoglobulin VH gene promoter that includes a functional consensus sequence. Cell. 1985 Jun;41(2):479–487. doi: 10.1016/s0092-8674(85)80021-0. [DOI] [PubMed] [Google Scholar]
  34. Muller-Sieburg C. E., Whitlock C. A., Weissman I. L. Isolation of two early B lymphocyte progenitors from mouse marrow: a committed pre-pre-B cell and a clonogenic Thy-1-lo hematopoietic stem cell. Cell. 1986 Feb 28;44(4):653–662. doi: 10.1016/0092-8674(86)90274-6. [DOI] [PubMed] [Google Scholar]
  35. Mulligan R. C., Berg P. Expression of a bacterial gene in mammalian cells. Science. 1980 Sep 19;209(4463):1422–1427. doi: 10.1126/science.6251549. [DOI] [PubMed] [Google Scholar]
  36. Neumann E., Schaefer-Ridder M., Wang Y., Hofschneider P. H. Gene transfer into mouse lyoma cells by electroporation in high electric fields. EMBO J. 1982;1(7):841–845. doi: 10.1002/j.1460-2075.1982.tb01257.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Nussenzweig M. C., Shaw A. C., Sinn E., Danner D. B., Holmes K. L., Morse H. C., 3rd, Leder P. Allelic exclusion in transgenic mice that express the membrane form of immunoglobulin mu. Science. 1987 May 15;236(4803):816–819. doi: 10.1126/science.3107126. [DOI] [PubMed] [Google Scholar]
  38. Ohashi P. S., Mak T. W., Van den Elsen P., Yanagi Y., Yoshikai Y., Calman A. F., Terhorst C., Stobo J. D., Weiss A. Reconstitution of an active surface T3/T-cell antigen receptor by DNA transfer. Nature. 1985 Aug 15;316(6029):606–609. doi: 10.1038/316606a0. [DOI] [PubMed] [Google Scholar]
  39. Pardoll D. M., Fowlkes B. J., Bluestone J. A., Kruisbeek A., Maloy W. L., Coligan J. E., Schwartz R. H. Differential expression of two distinct T-cell receptors during thymocyte development. Nature. 1987 Mar 5;326(6108):79–81. doi: 10.1038/326079a0. [DOI] [PubMed] [Google Scholar]
  40. Pardoll D. M., Fowlkes B. J., Lechler R. I., Germain R. N., Schwartz R. H. Early genetic events in T cell development analyzed by in situ hybridization. J Exp Med. 1987 Jun 1;165(6):1624–1638. doi: 10.1084/jem.165.6.1624. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Raulet D. H., Garman R. D., Saito H., Tonegawa S. Developmental regulation of T-cell receptor gene expression. Nature. 1985 Mar 7;314(6006):103–107. doi: 10.1038/314103a0. [DOI] [PubMed] [Google Scholar]
  42. Ritchie K. A., Brinster R. L., Storb U. Allelic exclusion and control of endogenous immunoglobulin gene rearrangement in kappa transgenic mice. Nature. 1984 Dec 6;312(5994):517–520. doi: 10.1038/312517a0. [DOI] [PubMed] [Google Scholar]
  43. Rusconi S., Köhler G. Transmission and expression of a specific pair of rearranged immunoglobulin mu and kappa genes in a transgenic mouse line. 1985 Mar 28-Apr 3Nature. 314(6009):330–334. doi: 10.1038/314330a0. [DOI] [PubMed] [Google Scholar]
  44. 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]
  45. Samelson L. E., Germain R. N., Schwartz R. H. Monoclonal antibodies against the antigen receptor on a cloned T-cell hybrid. Proc Natl Acad Sci U S A. 1983 Nov;80(22):6972–6976. doi: 10.1073/pnas.80.22.6972. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. Samelson L. E., Lindsten T., Fowlkes B. J., van den Elsen P., Terhorst C., Davis M. M., Germain R. N., Schwartz R. H. Expression of genes of the T-cell antigen receptor complex in precursor thymocytes. 1985 Jun 27-Jul 3Nature. 315(6022):765–768. doi: 10.1038/315765a0. [DOI] [PubMed] [Google Scholar]
  47. Snodgrass H. R., Dembić Z., Steinmetz M., von Boehmer H. Expression of T-cell antigen receptor genes during fetal development in the thymus. Nature. 1985 May 16;315(6016):232–233. doi: 10.1038/315232a0. [DOI] [PubMed] [Google Scholar]
  48. Snodgrass H. R., Kisielow P., Kiefer M., Steinmetz M., von Boehmer H. Ontogeny of the T-cell antigen receptor within the thymus. Nature. 1985 Feb 14;313(6003):592–595. doi: 10.1038/313592a0. [DOI] [PubMed] [Google Scholar]
  49. Staerz U. D., Pasternack M. S., Klein J. R., Benedetto J. D., Bevan M. J. Monoclonal antibodies specific for a murine cytotoxic T-lymphocyte clone. Proc Natl Acad Sci U S A. 1984 Mar;81(6):1799–1803. doi: 10.1073/pnas.81.6.1799. [DOI] [PMC free article] [PubMed] [Google Scholar]
  50. Storb U., Pinkert C., Arp B., Engler P., Gollahon K., Manz J., Brady W., Brinster R. L. Transgenic mice with mu and kappa genes encoding antiphosphorylcholine antibodies. J Exp Med. 1986 Aug 1;164(2):627–641. doi: 10.1084/jem.164.2.627. [DOI] [PMC free article] [PubMed] [Google Scholar]
  51. Storb U. Transgenic mice with immunoglobulin genes. Annu Rev Immunol. 1987;5:151–174. doi: 10.1146/annurev.iy.05.040187.001055. [DOI] [PubMed] [Google Scholar]
  52. Uematsu Y., Ryser S., Dembić Z., Borgulya P., Krimpenfort P., Berns A., von Boehmer H., Steinmetz M. In transgenic mice the introduced functional T cell receptor beta gene prevents expression of endogenous beta genes. Cell. 1988 Mar 25;52(6):831–841. doi: 10.1016/0092-8674(88)90425-4. [DOI] [PubMed] [Google Scholar]
  53. Wagner E. F., Stewart T. A., Mintz B. The human beta-globin gene and a functional viral thymidine kinase gene in developing mice. Proc Natl Acad Sci U S A. 1981 Aug;78(8):5016–5020. doi: 10.1073/pnas.78.8.5016. [DOI] [PMC free article] [PubMed] [Google Scholar]
  54. Wagner T. E., Hoppe P. C., Jollick J. D., Scholl D. R., Hodinka R. L., Gault J. B. Microinjection of a rabbit beta-globin gene into zygotes and its subsequent expression in adult mice and their offspring. Proc Natl Acad Sci U S A. 1981 Oct;78(10):6376–6380. doi: 10.1073/pnas.78.10.6376. [DOI] [PMC free article] [PubMed] [Google Scholar]
  55. Winoto A., Mjolsness S., Hood L. Genomic organization of the genes encoding mouse T-cell receptor alpha-chain. 1985 Aug 29-Sep 4Nature. 316(6031):832–836. doi: 10.1038/316832a0. [DOI] [PubMed] [Google Scholar]
  56. Zinn K., DiMaio D., Maniatis T. Identification of two distinct regulatory regions adjacent to the human beta-interferon gene. Cell. 1983 Oct;34(3):865–879. doi: 10.1016/0092-8674(83)90544-5. [DOI] [PubMed] [Google Scholar]

Articles from Molecular and Cellular Biology are provided here courtesy of Taylor & Francis

RESOURCES