Association of Sendai virion envelope and a mouse surface membrane polypeptide on newly infected cells: Lack of association with H-2K/D or alteration of viral immunogenicity

The recognition and lysis of virus-infected mouse cells by cytolytic T lymphocytes (CTL) requires that both viral and H-2 antigens be present on the target cell membrane. This recognition is known to be highly specific for both the viral and H-2K/D antigens. Thus, the targets must be matched with the CTL at either the H-2K and/or D loci. This phenomenon is referred to as H-2 restriction of CTL killing. One model that would explain this virus restriction postulates a CTL receptor that would recognize a new antigenic complex formed after some association of viral and H-2K/D or other host cell polypeptides on the target cell surface. To test for such associations, we have synthesized an 11 Å heterobifunctional cross-linking reagent possessing an amino active group at one terminus and a photoactivatable group at the other. This reagent (N-succinimidyl 4-azidophenyl-1,3'-dithiopropionate (SADP)) was coupled to purified Sendai virions by the amino-reactive end of the SADP molecule. This SADP-coupled virus was fused into the membranes of surface radioiodinated P815 cells, and target structures were allowed to form. Next, the photosensitive group on SADP was activated with ultraviolet light to covalently couple the viral proteins to any neighbouring cell surface proteins. The cellular neighbors were isolated from detergent extracts of membrane proteins after immunoprecipitation with antibody specific for Sendai virion proteins. The covalent cross-links between the nonradioactive Sendai proteins and the radioiodinated cellular polypeptide neighbors were broken, and the host cell polypeptides were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and detected by autoradiography. One of these neighboring cellular proteins had an apparent m.w. of 17,000, and none was found with the characteristic size and tryptic map of either the H-2K or D gene products. Thus, the H-2K or D proteins are unlikely to be SADP-detectable neighbors of Sendai viral antigens recognized by CTL. In further experiments, the complexes of Sendai virion proteins cross-linked to cellular polypeptide neighbors were isolated from the membrane of newly infected cells and were shown to be able to stimulate CTL in vitro with approximately the same efficiency as uncross-linked Sendai virion proteins. Thus, Sendai viral proteins in the membrane of newly infected cells do not appear to be in highly immunogenic complexes with either H-2K/D or any other cellular proteins.