The objective of the proposed study is the identification and characterization of viral gene products recognized by influenza A virus specific cytotoxic T lymphocytes (CTL). This work is of both practical and general importance since: 1) it may aid production of influenza vaccines which maximally prime for a CTL response, and 2) it will help define the requirements for CTL recognition of non-self antigens. The study will focus on CTL recognition of non-glycosylated viral proteins, one of which, the nucleoprotein (NP), is clealy shown to serve as a CTL target antigen by preliminary results. CTL recognition of NP and other non-glycosylated viral proteins will be examined using eukaryotic expression vectors cntaining cloned viral genes. Monoclonal antibodies will be used to aid in the biochemical and antigenic characterization of the cell surface forms of NP and other non-glycosylated proteins which are found to serve as CTL target antigens. The basis for the expression of these predominantly cytoplasmic proteins on cell surfaces and their recognition by CTL will be further examined using target cells sensitized for CTL lysis by acid mediated fusion of viral and cellular membranes. Also a focus of this proposal is examination of CTL cross-reactive recognition of influenza A virus glycoproteins derived from different subtypes. This will be investigated using eukaryotic expression vectors containing genes encoding viral glycoproteins. If CTL are found to recognize glycoproteins in a cross-reactive manner, attempts will be made to produce monoclonal antibodies with similar specificities. These antibodies will be characterized with respect to their biological activities, and the degree of overlap between their epitopes and the epitopes defined by CTL will be determined.
