title: Transfection of the CD8 alpha gene restores specific target cell lysis: factors that determine the function and the expression of CD8 in a cytotoxic T cell clone.
abstract: We investigated the functional role of CD8 and the control of CD8 expression in a constitutively activated murine cytotoxic T cell clone (C196), in CD8 deficient variants of this clone, and in cell lines derived by transfecting such variants with the CD8 alpha (Lyt-2) gene. CD8 deficient variants of C196 are deficient in specific target cell lysis but retain the ability to perform anti-CD3 induced cytolytic function. Following transfection with the Lyt-2 gene, specific target cell lysis was restored in some but not all CD8 positive transfectants whereas no alteration of anti-CD3 induced lysis was observed. All CD8 deficient variants studied lost the surface expression of both Lyt-2 and Lyt-3 polypeptide chains. However, while Lyt-2 mRNA was abolished as well, typical CD8 deficient variants retained wild type levels at Lyt-3 mRNA. Moreover, a low level of cytoplasmic Lyt-3 protein was demonstrable. Following transfection with the Lyt-2 gene, both Lyt-2 and Lyt-3 polypeptide chains reappeared on the membrane. In one atypical CD8 deficient variant that carries a mutated Lyt-3 gene and fails to express Lyt-3 mRNA, Lyt-2 transfection causes membrane-reappearance of Lyt-2 only. These results may reflect the occurrence on normal T cells of Lyt-2/Lyt-3 heterodimers and of Lyt-2/Lyt-2 homodimers, whereas surface expression of Lyt-3 alone has not been observed. In CD8 deficient variant, a particular restriction site 5' at the Lyt-2 gene is methylated which is undermethylated in the wild type C196. Culture of such variants in 5-azacytidine partially restored CD8 expression. This suggests a negative correlation of Lyt-2 transcription with site-specific DNA methylation in the C196 system. However, results on T cells unrelated to C196 suggests that the site whose methylation appears to be critical in C196 is not responsible for Lyt-2 transcription in all T cells.
authors: Max-Planck-Institut für Immunobiologie, Freiburg, FRG.
authors: R
authors: R
authors: Schmidt-Ullrich
authors: K
authors: K
authors: Eichmann
mesh_terms: D000818:Animals; D000945:Antigens, Differentiation, T-Lymphocyte / Q000235:genetics* / Q000502:physiology; D000950:Antigens, Ly / Q000235:genetics; D016827:CD8 Antigens; D002460:Cell Line; D003602:Cytotoxicity, Immunologic; D005786:Gene Expression Regulation; D007694:Killer Cells, Natural / Q000276:immunology; D051379:Mice; D011994:Recombinant Proteins / Q000502:physiology; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology*; D014162:Transfection
chemical_list: D000945:Antigens, Differentiation, T-Lymphocyte; D000950:Antigens, Ly; D016827:CD8 Antigens; D011994:Recombinant Proteins