Proper functioning of the immune system, in detecting and mounting an immune response against invading pathogens, is dependent on lymphocyte recirculation through secondary lymphoid organs. Lymphocyte entry and exit into these lymphoid tissues is a tightly regulated process. Disruption of this process can lead to sequestration of lymphocytes in lymphoid organs, as seen in infection with human immunodeficiency virus, and may contribute to immunosuppression of the host. A variety of cytokines, including interferon-a/? (IFN- a/? are known to transiently block lymphocyte egress. Since an increasing number of cytokines, including IFN-a/b are used as therapeutic agents, it is essential to understand the molecular mechanisms by which they affect lymphocyte trafficking. We have found that treatment of mice with the IFN-a/? inducer, polyinosine-polycytosine (PIC), inhibited egress through both lymphocyte intrinsic and extrinsic mechanisms. Lymphocyte egress is dependent on sphingosine-1 -phosphate receptor-1 (S1P1), and IFN-a/b was found to inhibit lymphocyte responsiveness to its ligand, sphingosine-1-phosphate (S1P). Loss of lymphocyte responsiveness to S1P was mediated by the transmembrane C-type lectin, CD69. In co-expression studies, CD69 co-immunoprecipitated with S1P1. Furthermore, CD69 inhibited S1P1 chemotactic function and led to S1P1 down-modulation. This is the first demonstration of negative regulation of a G-protein coupled receptor by a type II transmembrane glycoprotein. The goal of this application is to further understand the effects of IFN-a/? on lymphocyte egress with emphasis on the molecular basis of regulation of S1P1 by CD69. Cellular and biochemical approaches will be utilized to define (1) the mechanism of lymphocyte extrinsic effects of IFN-a/b on exit; (2) molecular requirements for S1P-I-CD69 interaction; (3) the fate of the molecular complex; and (4) effects on signaling pathways downstream of S1P1 . The candidate is a physician-scientist who is proposing a 3-year mentored research experience with Dr. Arthur Weiss at the University of California, San Francisco. The proposed training program is designed with the goal of preparing the applicant to establish an independent laboratory in an academic department. ? ? ?
