Defining mechanisms of KSHV pathogenesis using MHV68-KSHV chimeric viruses

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PROJECT SUMMARY Gammaherpesviruses (GHVs) are DNA tumor viruses that establish lifelong, chronic infections of host lymphocytes. The expression of viral gene products that manipulate host cell physiology and thwart antiviral immune responses places the infected host at risk for numerous cancers. For individuals with AIDS, infection by Kaposi sarcoma-associated herpesvirus (KSHV) is a major cause of morbidity and mortality. However, KSHV does not readily infect mice, which complicates attempts to define mechanisms by which KSHV establishes long- term infections and disease. To overcome this barrier, we have used a chimeric virus approach in which KSHV genes are transferred into the closely related virus, murine gammaherpesvirus 68 (MHV68), which is a natural rodent pathogen that readily infects laboratory mice. We used this MHV68-KSHV chimeric virus approach to evaluate the KSHV latency-associated nuclear antigen (kLANA) during productive viral replication, latency establishment, and maintenance. We found that kLANA was sufficient to replace MHV68 LANA (mLANA) for viral latency. However, kLANA, but not mLANA, suppressed MHV68 lytic replication by inhibiting the activity of the promoter for lytic transactivator RTA. Our work demonstrates that chimeric viruses offer opportunities for dissecting functions of KSHV proteins in a living host, and that this system can define evolutionarily acquired features of a viral oncoprotein. Experiments described in this proposal will further define functions of KSHV LANA in viral pathogenesis and evolutionary divergence from mLANA. We will use the repressed lytic replication of the kLANA-expressing chimeric virus to define how the amplitude of lytic replication influences immunity to infection and oncogenic potential of the virus. Finally, we will extend our chimeric virus studies with LANA, dovetailing them with MHV68-KSHV chimeric viruses for v-Cyclin and v-GPCR, to develop the next generation of chimeras in which multiple MHV68 genes are replaced with their KSHV counterparts. Through this work we will improve small animal models to enable molecular mechanistic studies of KSHV oncoproteins in viral infection and disease and provide a preclinical testing platform to foster the development of therapies that target LANA and other viral proteins to treat or prevent KSHV-related cancers.