GENETICS AND FUNCTIONS OF KSHV GLYCOPROTEINS GM AND GN

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This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Kaposi's sarcoma-associated herpesvirus (KSHV), or human herpesvirus 8, is a lymphotropic oncogenic virus that has been implicated in the pathogenesis of Kaposi's sarcoma; body cavity-based B-cell lymphoma (BCBL), or primary effusion lymphoma; and some forms of multicentric Castleman's disease. We have developed a conditional silencing system for partial inhibition of KSHV viral glycoprotein synthesis using specific siRNAs. In this system, protein synthesis can be reconstituted using codon-optimized genes that are not susceptible to siRNA inhibition. We have succesfully used this system to demonstrate that KSHV glycoprotein B (gB) is necessary for virion egress from BCBL-1 cells and virus infectivity (Subramanian, D'Auvergne, et al. J. Virol. 2008, 82:7144-54). To investigate the potential role of gB in tumorigenesis, BCBL-1 cells transiently transfected with anti-gB siRNAs and codon optimized gB were mixed with matrigel and injected subcutaneously in nude mice. Direct measurement of tumors revealed that BCBL-1 cells transfected with anti-gB siRNAs produced tumors significantly smaller than mock-transfected BCBL-1 cells. Co-transfection of codon optimized gB appeared to abrogate the inhibition of tumor formation by siRNAs. These results show that gB is important for infectivity, virion egress and pleural effusion lymphoma (PEL) formation in mice. As part of a graduate student project, we have also worked on the effect of polycations on facilitating KSHV virus entry into 293 cells. It was found that DEAE-dextran, poly-lysine, polybrene, protamine and protamine sulfate enhanced the KSHV entry into 293 and other cell lines when added at the time of infection.