) Herpesviruses are associated with a variety of human malignancies, but the molecular mechanisms of cell transformation and tumorigenesis are not known. Immunosuppressed AIDS patients are particularly prone to the complications of herpesvirus-induced tumors. A new human herpesvirus, HHV8, was recently identified in association with Kaposi's sarcoma (KS) in AIDS and other patients. Through sequence analysis the applicant has identified a homolog of the human bcl-2 oncogene-encoded by HHV8. The bcl-2 oncogene was identified at translocation breakpoints characteristic of follicular B cell lymphomas. Bcl-2 contributes to tumorigenesis because it is a potent inhibitor of programmed cell death and can function in a wide variety of cell types that are induced to undergo apoptosis by a broad range of stimuli. However, the mechanism by which bcl-2 blocks cell death is not known. The HHV-8 homolog, designated KSbcl-2, also has anti-apoptotic activity and may cooperate with other factors to facilitate transformation similar to that observed for adenovirus E1b. Preliminary studies on the bcl-2 homolog encoded by herpesvirus saimiri indicate that gamma herpesviruses require an inhibitor of apoptosis to successfully complete the lytic replication cycle, suggesting, at the least, that a herpesvirus bcl-2 homolog facilitates transformation by supporting viral replication. (i) To study the bcl-2 homolog of HHV-8, the applicant will identify and map the KSbcl-2 mRNAs in cultured cells and determine if they are expressed during the lytic cycle, the latent cycle, or both. An antibody generated to KSbcl-2 will be used to confirm protein expression during different phases of the virus life cycle in cultured cells and to determine if KSbcl-2 is expressed in KS tumors. (ii) Heterodimerization by bcl-2 family members regulates their function. Therefore, the applicant will determine which cellular bcl-2 family members interact with KSbcl-2 and how these interactions affect the function of KSbcl-2. (iii) The role of KSbcl-2 in virus infection will be studied by determining if KSBCL-2 can rescue the null plaque phenotype of bcl-2-deficient herpes saimiri. (iv) To further understand the function of KSbcl-2, the applicant seeks to identify cellular proteins that mediate the oncogenic/anti-apoptotic potential of KSbcl-2 using the yeast two-hybrid interactive cloning strategy and direct biochemical approaches.
