The long-term objective of the proposed research is to determine the structure and function of lymphoid cell surface proteins that may play important roles in the lytic activity of murine natural killer (NK) cells. We have identified a cDNA clone that directs the expression of a murine lymphoid cell surface antigen which is recognized by the monoclonal antibody (mAb) A1 and which is expressed by rare splenocytes. This probe hybridizes to multiple bands on Southern blots, consistent with the hypothesis that there are several other, highly related genes. Our preliminary data demonstrate that the A1 multigene family maps to a contiguous stretch of mouse chromosome 6 to which another lymphoid cell surface antigen, NK1.1, also maps. The NK1.1 antigen is the most specific marker of murine NK cells, and since the NK1.1 antigen is also coordinately expressed with the A1 antigen, the NK1.1 antigen may be a member of the A1 multigene family. Moreover, the A1 and NK1.1 antigens may be involved in Nk cell effector function. The first specific aims of this project is to characterize the NK1.1 antigen and other members of the A1 multigene family at the molecular level. If the NK1.1 and A1 antigens are molecularly similar, we will exploit the similarity to clone and determine the primary structure of the NK1.1 antigen. This approach should also result in the cloning of other members of the A1 multigene fAmily. If the NK1.1 antigen is not homologous to the A1 antigen, we will clone the NK1.1 gene by immunoselection in an eukaryotic expression system. The second specific aim is to express newly cloned members of the A1 gene family. We will use newly cloned cDNAs to identify cells and tissues that express these antigens. We also will stably express these cDNAs in transfected eukaryotic cells in order analyze function. These cells will be employed also to derive new mAbs that in turn will be useful for other functional analyses. The final specific aim is to analyze the functional role of the NK1.1 and A1 antigens in NK-mediated lysis of target cells. We will test the hypothesis that these antigens are involved in NK effector function by assaying the ability of cytotoxic T lymphocyte (CTL) clones, stably transfected with the A1 and NK1.1 antigens, to lyse normal targets of Nk cells, and by determining whether mAbs against these antigens block or enhance NK-mediated cell lysis. Results of these experiments should enhance our understanding of the roles played by members of the A1 multigene family and/or the NK1.1 antigen in the recognition and killing of specific targets by murine NK cells.
