Adeno-associated virus (AAV) is a nonpathogenic human parvovirus with a unique survival strategy which includes helper virus dependency and frequent long term stable provirus integration. It is well documented that AAV encodes a function with significant anti-proliferative properties. Consistent with these studies are seroepidemiologic studies which show that AAV infection is negatively associated with human genital cancer. In three studies this P.I. has mapped the anti-proliferation effect to the full length AAV Rep78 gene using BPV and c-EJ-H-ras as target oncogenes. Rep78 has been described as a viral """"""""antioncogene"""""""" because of similarities with p105-RB and p53. Rep78 is also involved in AAV DNA replication and transcriptional regulation, and these same biochemical properties of Rep78 may also be involved in its anti- proliferative activity. It is the objective of this proposal to understand the role of Rep78 anti-proliferative function in AAV biology and heterologous oncogene regulation. To do so, it is proposed to structurally analyze and compare the domains of the Rep78 molecule which are needed for antioncogene and AAV directed functions. Furthermore, to address the other half of the interaction, it is proposed to analyze the level of heterologous oncogene (c-H-ras) regulation and to define the cis target sequences through which Rep78 inhibits.
Specific Aim #1 : Study the effector. Map the domains of the Rep78 protein required for anti-proliferation activity (inhibition of c-EJ-H- ras), AAV DNA replication, and AAV transcriptional transactivation.
Specific Aim #2 : Study the target. Investigate the level at which inhibition of c-H-ras takes place and identify the cis c-H-ras regulatory sequences which are required for Rep78 induced inhibition.
