Kaposi's Sarcoma herpesvirus (KSHV or HHV-8) is the causative agent of angiogenic skin malignancies that occur primarily in association with the AIDS epidemic. Despite success in controlling HIV levels in Western countries with HAART, affected patients are rarely cured of their malignancies. Furthermore, because of the spread of HIV, Kaposi's Sarcoma has now become the most prevalent tumor encountered in Southern Africa. The presence of Kaposi's Sarcoma-Associated Herpesvirus in KS, and both Epstein-Barr virus (EBV) and KSHV in Primary Effusion Lymphomas (PELs) provides an important target for the investigation of the pathogenesis and ultimately the specific treatment of these neoplasms. The overall goal of this Program Project is to investigate how KSHV takes control of vascular endothelial cells and B-lymphocytes, with a mechanistic focus on specific viral genes expected to be relevant to tumorigenesis, prevention, and treatment of KS and PEL. In particular, this PPG concentrates on evaluating and using a cell culture model for KS involving primary adult human microvascular endothelial cells (DMVEC), in which both an efficient and stable KSHV latent state and lytic cycle reactivation occur in converted KS-like spindle cells. Current models of KSHV pathogenesis envisage roles for the combined activities of both constitutively expressed KSHV latency gene products plus the occasional expression of abortive lytic cycle gene products leading to long-term host cell alterations. In this revised version of the project, Project 1 (Gary Hayward) will focus on control of transcription and post-translational events during maintenance of latency in KSHV-infected DMVEC, including changes in key EC protein expression and function in the infected spindle cells, and the biological roles of the viral vMIR1,vMIR2 and vFLIP proteins. Project 2 (S. Diane Hayward and Richard Ambinder) will evaluate cellular protein interactions and repression of cellular gene expression by the LANA1 protein involved in KSHV genome maintenance and in driving cell proliferation. Project 3 (John Nicholas) targets two KSHV homologues of cellular beta-chemokines which have anti-apoptotic functions and induce VEGF, and include autocrine interactions with vGPCR that may play a role in supporting neoplastic proliferation and lytic reactivation. These three cooperative Projects all address overlapping aspects of KSHV pathogenesis, focusing on the role of viral and cellular genes in maintenance of latency and the role of the early lytic cycle captured cellular genes. The PPG is centered around extensive utilization of the expertise and reagents generated by the skilled personnel in two Laboratory Cores involving KSHV BAG mutants and genetics, novel lentivirus and adenovirus vectors for expression of KSHV genes and siRNAs, and improved specific procedures for analysis of gene expression by custom gene array, IFA and IHC, which greatly enhance synergy, reagent sharing and overall beneficial interactions between all of the Projects.
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