Die Infektion endothelialer Zellen mit dem humanen Herpesvirus 8 (KSHV/HHV8): Mechanismen der Persistenz, Replikation und abnormaler Differenzierung

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Kaposi's Sarcoma associated herpesvirus (KSHV/HHV-8), is an essential factor in the pathogenesis of Kaposi's sarcoma (KS), a tumour derived from endothelial cells. KSHV infects, and probably persists in, endothelial cells and induces their atypical differentiation into KS spindle cells, the hallmark of this tumour. Viral proteins expressed during latent persistence and lytic replication may contribute to the development KS spindle cells. However, in characterising the function of individual viral proteins, all previous studies have studied them in isolation, outside the context of the remainder of the virus genome, and mostly after overexpression. The experience with other herpesviruses suggests that this may lead to misleading interpretations. The lack of an efficient in vitro culture system has so far precluded experiments with KSHV mutants. Given the recent improvements in cell culture systems for KSHV we propose to construct a recombinant molecular clone of KSHV in a 'bacterial artificial chromosome' (BAC) and to use it for the generation of a mutant transposon library as well as mutants of individually targeted genes. The aim of the proposal is to identify KSHV mutants that have lost the ability to replicate and/or persist in endothelial cells and to induce spindle cell formation in primary endothelial cells. This will allow the identification of viral genes that are essential for KSHV-mediated pathogenesis in endothelial cells. The results will contribute to our understanding of how herpesviruses in general infect and persist in endothelial cells.