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. 1988 May;64(1):129–134.

Appearance of granule-associated molecules during activation of cytolytic T-lymphocyte precursors by defined stimuli.

J A Garcia-Sanz 1, F Velotti 1, H R MacDonald 1, D Masson 1, J Tschopp 1, M Nabholz 1
PMCID: PMC1385197  PMID: 3260214

Abstract

Lysis of target cells by cytolytic T lymphocytes (CTL) is associated with the exocytosis of cytoplasmic granules. Purified granules from CTL cell lines contain a pore-forming protein (perforin), tree serine esterases, granzyme A (60,000 MW), granzyme B (29,000 MW), and granzyme C (27,000 MW). We have compared the kinetics of appearance of cytolytic activity with that of perforin and granzyme A activity during activation of lymphocytes from normal animals with leukoagglutinin (LA) and recombinant interleukin-2 (rIL-2). Unstimulated lymph node cells do not express any of these activities, which appear between Day 3 and Day 4 of stimulation and increase rapidly to reach a pronounced peak on Day 6. On Day 7 all the activities are considerably lower, even though the cells still proliferate exponentially. There is a good correlation between the kinetics of appearance of all of these activities. Using antisera against perforin and against granzyme C, one can detect positive cytoplasmic granules in a small fraction of cells on Day 3; by Day 5, 80-90% of the cells are stained. This proportion decreases again on Day 7.

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Selected References

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

  1. Bevan M. J., Langman R. E., Cohn M. H-2 antigen-specific cytotoxic T cells induced by concanavalin A: estimation of their relative frequency. Eur J Immunol. 1976 Mar;6(3):150–156. doi: 10.1002/eji.1830060303. [DOI] [PubMed] [Google Scholar]
  2. Brunet J. F., Dosseto M., Denizot F., Mattei M. G., Clark W. R., Haqqi T. M., Ferrier P., Nabholz M., Schmitt-Verhulst A. M., Luciani M. F. The inducible cytotoxic T-lymphocyte-associated gene transcript CTLA-1 sequence and gene localization to mouse chromosome 14. Nature. 1986 Jul 17;322(6076):268–271. doi: 10.1038/322268a0. [DOI] [PubMed] [Google Scholar]
  3. Devos R., Plaetinck G., Cheroutre H., Simons G., Degrave W., Tavernier J., Remaut E., Fiers W. Molecular cloning of human interleukin 2 cDNA and its expression in E. coli. Nucleic Acids Res. 1983 Jul 11;11(13):4307–4323. doi: 10.1093/nar/11.13.4307. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Erard F., Corthesy P., Nabholz M., Lowenthal J. W., Zaech P., Plaetinck G., MacDonald H. R. Interleukin 2 is both necessary and sufficient for the growth and differentiation of lectin-stimulated cytolytic T lymphocyte precursors. J Immunol. 1985 Mar;134(3):1644–1652. [PubMed] [Google Scholar]
  5. Erard F., Nabholz M., Dupuy-D'Angeac A., MacDonald H. R. Differential requirements for the induction of interleukin 2 responsiveness in L3T4+ and Lyt-2+ T cell subsets. J Exp Med. 1985 Nov 1;162(5):1738–1743. doi: 10.1084/jem.162.5.1738. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Erard F., Nabholz M., MacDonald H. R. Antigen stimulation of cytolytic T lymphocyte precursors: minimal requirements for growth and acquisition of cytolytic activity. Eur J Immunol. 1985 Aug;15(8):798–803. doi: 10.1002/eji.1830150811. [DOI] [PubMed] [Google Scholar]
  7. Garcia-Sanz J. A., Plaetinck G., Velotti F., Masson D., Tschopp J., MacDonald H. R., Nabholz M. Perforin is present only in normal activated Lyt2+ T lymphocytes and not in L3T4+ cells, but the serine protease granzyme A is made by both subsets. EMBO J. 1987 Apr;6(4):933–938. doi: 10.1002/j.1460-2075.1987.tb04841.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Gershenfeld H. K., Weissman I. L. Cloning of a cDNA for a T cell-specific serine protease from a cytotoxic T lymphocyte. Science. 1986 May 16;232(4752):854–858. doi: 10.1126/science.2422755. [DOI] [PubMed] [Google Scholar]
  9. Guesdon J. L., Ternynck T., Avrameas S. The use of avidin-biotin interaction in immunoenzymatic techniques. J Histochem Cytochem. 1979 Aug;27(8):1131–1139. doi: 10.1177/27.8.90074. [DOI] [PubMed] [Google Scholar]
  10. Hardt C., Sato N., Wagner H. Functional and biochemical characteristics of a murine interleukin 2 receptor-inducing factor. Eur J Immunol. 1987 Feb;17(2):209–216. doi: 10.1002/eji.1830170210. [DOI] [PubMed] [Google Scholar]
  11. Henkart P. A. Mechanism of lymphocyte-mediated cytotoxicity. Annu Rev Immunol. 1985;3:31–58. doi: 10.1146/annurev.iy.03.040185.000335. [DOI] [PubMed] [Google Scholar]
  12. Jenne D., Rey C., Masson D., Stanley K. K., Herz J., Plaetinck G., Tschopp J. cDNA cloning of granzyme C, a granule-associated serine protease of cytolytic T lymphocytes. J Immunol. 1988 Jan 1;140(1):318–323. [PubMed] [Google Scholar]
  13. Kramer M. D., Binninger L., Schirrmacher V., Moll H., Prester M., Nerz G., Simon M. M. Characterization and isolation of a trypsin-like serine protease from a long-term culture cytolytic T cell line and its expression by functionally distinct T cells. J Immunol. 1986 Jun 15;136(12):4644–4651. [PubMed] [Google Scholar]
  14. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  15. Lobe C. G., Havele C., Bleackley R. C. Cloning of two genes that are specifically expressed in activated cytotoxic T lymphocytes. Proc Natl Acad Sci U S A. 1986 Mar;83(5):1448–1452. doi: 10.1073/pnas.83.5.1448. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Martin D. E., Zalman L. S., Jung G., Müller-Eberhard H. J. Induction of synthesis of the cytolytic C9 (ninth component of complement)-related protein in human peripheral mononuclear cells by monoclonal antibody OKT3 or interleukin 2: correlation with cytotoxicity and lymphocyte phenotype. Proc Natl Acad Sci U S A. 1987 May;84(9):2946–2950. doi: 10.1073/pnas.84.9.2946. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Masson D., Corthésy P., Nabholz M., Tschopp J. Appearance of cytolytic granules upon induction of cytolytic activity in CTL-hybrids. EMBO J. 1985 Oct;4(10):2533–2538. doi: 10.1002/j.1460-2075.1985.tb03967.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Masson D., Nabholz M., Estrade C., Tschopp J. Granules of cytolytic T-lymphocytes contain two serine esterases. EMBO J. 1986 Jul;5(7):1595–1600. doi: 10.1002/j.1460-2075.1986.tb04401.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Masson D., Tschopp J. A family of serine esterases in lytic granules of cytolytic T lymphocytes. Cell. 1987 Jun 5;49(5):679–685. doi: 10.1016/0092-8674(87)90544-7. [DOI] [PubMed] [Google Scholar]
  20. Masson D., Tschopp J. Isolation of a lytic, pore-forming protein (perforin) from cytolytic T-lymphocytes. J Biol Chem. 1985 Aug 5;260(16):9069–9072. [PubMed] [Google Scholar]
  21. Pasternack M. S., Eisen H. N. A novel serine esterase expressed by cytotoxic T lymphocytes. 1985 Apr 25-May 1Nature. 314(6013):743–745. doi: 10.1038/314743a0. [DOI] [PubMed] [Google Scholar]
  22. Pasternack M. S., Verret C. R., Liu M. A., Eisen H. N. Serine esterase in cytolytic T lymphocytes. Nature. 1986 Aug 21;322(6081):740–743. doi: 10.1038/322740a0. [DOI] [PubMed] [Google Scholar]
  23. Podack E. R., Konigsberg P. J. Cytolytic T cell granules. Isolation, structural, biochemical, and functional characterization. J Exp Med. 1984 Sep 1;160(3):695–710. doi: 10.1084/jem.160.3.695. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Podack E. R., Young J. D., Cohn Z. A. Isolation and biochemical and functional characterization of perforin 1 from cytolytic T-cell granules. Proc Natl Acad Sci U S A. 1985 Dec;82(24):8629–8633. doi: 10.1073/pnas.82.24.8629. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Velotti F., MacDonald H. R., Nabholz M. Granzyme A secretion by normal activated Lyt-2+ and L3T4+ T cells in response to antigenic stimulation. Eur J Immunol. 1987 Aug;17(8):1095–1099. doi: 10.1002/eji.1830170804. [DOI] [PubMed] [Google Scholar]
  26. Young J. D., Leong L. G., Liu C. C., Damiano A., Cohn Z. A. Extracellular release of lymphocyte cytolytic pore-forming protein (perforin) after ionophore stimulation. Proc Natl Acad Sci U S A. 1986 Aug;83(15):5668–5672. doi: 10.1073/pnas.83.15.5668. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Young J. D., Leong L. G., Liu C. C., Damiano A., Wall D. A., Cohn Z. A. Isolation and characterization of a serine esterase from cytolytic T cell granules. Cell. 1986 Oct 24;47(2):183–194. doi: 10.1016/0092-8674(86)90441-1. [DOI] [PubMed] [Google Scholar]

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