Human herpesvirus-8 (HHV-8, also called KSHV) encodes several proteins that have been implicated in virus-associated pathogenesis. The viral interleukin-6 (vlL-6) and chemokine receptor (vGPCR) have been studied extensively due to their mitogenic and angiogenic properties, their promotion of tumor growth in in vitro and in vivo experimental systems, the proliferative effects of IL-6 signalling on Kaposi's sarcoma (KS), myeloma and primary effusion lymphoma (PEL) cell growth, and the ability of vGPCR to induce KS-like disease in receptor-transduced mice. However, angiogenic activities are also effected by each of the vchemokines (vCCL-1, vCCL-2, vCCL-3), and we have determined that vCCL-1 and vCCL-2 specify antiapoptotic activities, as measured in HHV-8 latently-infected PEL cells challenged with dexamethasone or serum withdrawal. These activities, coupled with the ability of vCCL-2 to regulate the signalling by vGPCR, suggest strongly that the v-chemokines play direct, autocrine roles in lytic replication. We hypothesize that these viral ligands serve to prolong survival of infected cells during lytic replication and to promote intracellular conditions that favor virus production. The anti-apoptotic functions of vCCL-1 and vCCL-2, like similar activities of their cellular counterparts, have not been studied in detail, and the roles of the ligands in virus productive replication have not been investigated. The purpose of this application is to build on our previous work on v-chemokine function and vGPCR signal transduction to investigate the roles of the vCCL- 1 and vCCL-2 in lytic replication and the relevance of vCCL-2:vGPCR interactions to replication efficiency.
The specific aims are: (1) to determine the mechanisms of v-chemokine anti-apoptotic signalling, (2) to map the ligand and receptor residues involved in antagonistic vCCL-2:vGPCR interactions, and (3) to determine the roles of the v-chemokines in lytic replication through the generation and utilization in replicationpermissive endothelial cell culture systems of vCCL- and vGPCR-altered HHV-8 recombinant viruses, and by using ribozymes or siRNAs to deplete vCCLs during lytic reactivation in PEL cells. The data from these studies will characterize the roles of the v-chemokines in virus replication and may provide the basis for the development of anti-viral vCCL-2-based vGPCR antagonists.
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