A major question in the pathogenesis of the Autoimmune Deficiency Syndrome (AIDS) concerns the mechanism by which uninfected CD4+ T cells become depleted in peripheral blood. HIV gp120 binding to CD4 of uninfected cells may generate intracellular signals that induce non- responsiveness (anergy), cell death (apoptosis) or syncytia formation. In this grant proposal, we report the identification of two lipid kinases, phosphotidylinositol-3 kinase (PI-3 kinase) and phosphotidylinositol-4 kinase (PI-4 kinase) associated with the CD4-lck complex. Of great interest, anti-HIV-1 gp120 binding to CD4 significantly increased levels of precipitable PI-3 and PI-4 kinase activity. To our knowledge, this represents the first case of a CD4 associated second messenger whose presence and/or activity is altered by gp120 binding. Of particular interest, PI-3 kinase has been found essential to signalling via other receptors, and is structurally-related to the yeast Vps 34 protein which controls membrane sorting and fusion. We propose that HIV-1 may utilize PI-3/PI-4 kinases in the generation of signals that produce immune defects. to this end, we plan to examine the biochemical basis of the interaction (i.e., stoichiometry, assembly, folding of the kinase), its regulation by HIV-gp120 (i.e., receptor aggregation phosphorylation) and to determine the binding sites between p56lck and PI-3/PI-4 kinase. Preliminary data using GST-fusion proteins indicate that lipid kinases bind to the SH2 region of lck. Using association-defective versions of the kinases, we will attempt in transfection studies to reverse HIV gp120 impaired signalling and/or syncytium formation. This proposal should allow elucidation of the possible role played by CD4 coupled PI-3/PI-4 kinase in HIV-induced immune defects.
| Rudd, C E; Janssen, O; Cai, Y C et al. (1994) Two-step TCR zeta/CD3-CD4 and CD28 signaling in T cells: SH2/SH3 domains, protein-tyrosine and lipid kinases. Immunol Today 15:225-34 |
