Studies on the induction and expression of T-cell-mediated immunity. XIII. Membrane-associated antigens of cytotoxic T lymphocytes involved in cytotoxicity.
An xenogeneic rat anti-mouse T-cell serum, designated RAT*, has been shown to block the cytolytic activity of cytotoxic T lymphocytes (CTL) at a postbinding step. RAT* serum or the IgG fraction was extensively absorbed with the target cell, P815, a DBA mastocytoma, and used with or without further absorption to immunoprecipitate specific molecules from radiolabeled membrane extracts of CTL derived from either in vivo-allosensitized mice or from cytotoxic clones maintained in in vitro cultures. Cell surface sialic acid residues were labeled by oxidation with sodium periodate (NaIO4) and reduction with tritiated sodium borohydride ([3H]NaBH4). Alternatively, cell surface proteins were labeled with 125I by lactoperoxidase-catalyzed iodination. Nonidet P-40 (NP-40)-solubilized radiolabeled membranes were then immunoprecipitated with RAT* serum and analyzed by sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis (PAGE). Three membrane-associated molecules of 95,000, 140,000 and 180,000 Mr were found by such analysis. The sensitivity of these three molecules to trypsinization and their susceptibility to labeling with [3H]NaBH4 suggested that they are glycoproteins. Moreover, when RAT* serum or the IgG fraction was absorbed with various cell types, its ability to immunoprecipitate the three molecules correlated with its ability to block cytolysis. Adsorption of RAT* serum with CTL, but not with nonimmune thymocytes, significantly reduced the ability of RAT* serum to inhibit cytotoxicity and to immunoprecipitate the 95k, 140k, and 180k molecules. Thus, these findings suggest that one or more of these cell surface molecules of CTL may be involved in the cytolytic process.
D000818:Animals; D000954:Antigens, Surface / Q000032:analysis*; D000961:Antilymphocyte Serum / Q000494:pharmacology; D001667:Binding, Competitive; D001894:Borohydrides / Q000378:metabolism; D011232:Chemical Precipitation; D003602:Cytotoxicity, Immunologic; D007111:Immunity, Cellular; D008297:Male; D008565:Membrane Proteins / Q000032:analysis*; D051379:Mice; D008810:Mice, Inbred C57BL; D008811:Mice, Inbred DBA; D008970:Molecular Weight; D051381:Rats; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology* / Q000378:metabolism
D000954:Antigens, Surface; D000961:Antilymphocyte Serum; D001894:Borohydrides; D008565:Membrane Proteins; C025364:sodium borohydride