Defining KSHV LANA functions in viral pathogenesis and immune evasion

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 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. The KSHV latency-associated nuclear antigen (kLANA) is an oncogene that modulates viral and host-cell transcription and is required for maintaining latent viral episomes as infected cells divide. We used this MHV68-KSHV chimeric virus approach to evaluate kLANA functions during productive viral replication, latency establishment, and maintenance. Our published work demonstrated that kLANA was sufficient to replace MHV68 LANA (mLANA) for viral latency in mice. Remarkably, kLANA, but not mLANA, suppressed MHV68 lytic replication by inhibiting the activity of the promoter for lytic transactivator RTA, suggesting that KSHV-specific LANA functions are also active in the chimeric MHV68. In new preliminary data we demonstrate that suppressed lytic replication by kLANA correlates with a drastic reduction in adaptive immunity to chimeric virus infection, which supports the guiding hypothesis for this proposal that kLANA- mediated inhibition of viral replication and reactivation is immune evasive. Experiments in Aim 1 will define the scope of immune avoidance by MHV68-KSHV chimeric virus and determine the impact of reduced immunity on outcomes of infection. We will simultaneously identify kLANA’s role as a transcription regulator in the process. Experiments in Aim 2 will leverage reduced replication and immune evasion to define how kLANA functions impact viral pathogenesis during immune impairment, especially when CD4 T cells are depleted as in non-drug- controlled HIV infection. We have also developed a combinatorial MHV68-KSHV chimeric virus that encodes multiple KSHV oncogenic latency genes for future development of chimeric virus models. Through the use of innovative and rigorously established small-animal models of infection and disease, we will better define mechanisms through which KSHV LANA contributes to latency and pathogenesis. This work will increase the experimental tractability and relevance of a preclinical model and may foster the development of therapies that target LANA and other viral proteins to treat or prevent KSHV-related cancers as a result.