Recent studies from the investigator's laboratory have identified a cellular nuclear protein, Sam68, which specifically interacts with RRE and can substitute for as well as synergize with Rev in RRE mediated gene expression and virus replication. The major goals of this proposal are to functionally characterize Sam68 protein in RRE-mediated transactivation and determine the relevance of Sam68 in HIV replication.
The specific aims are: 1) to functionally characterize the interactions of Sam68 with Rev and/or RRE RNA in RRE-mediated gene expression using truncated and site-directed mutants to map domains of Sam68 and its binding sites on RRE as well as Rev that are functionally involved in RRE-mediated gene expression and virus replication; 2) to investigate the functional relevance of Sam68 in HIV replication by inactivation of Sam68 genes using ribozymes, antisense, and anti- Sam68 antibodies.
This Aim will also generate an in vitro model for the enhancement of HIV replication in NIH 3T3 cells by Sam68; 3) to assess the long-term protection of primary cells expressing transdominant mutants of Sam68 from HIV infection by transducing T lymphocytes and primary cells with a retroviral vector encoding transdominant negative mutants of Sam68 and assessing long-term protection of these cells from HIV-1 infection. The effect of Sam68 on HIV mutants that are resistant to Rev M10 will also be assessed. These studies may allow the development of novel anti-viral agents.
