Project Summary Reactivation of HSV in latently infected sensory neurons is of considerable clinical importance especially with respect to eye disease. Development of effective alternative strategies to limit reactivation has been limited by an insufficient understanding of mechanisms that regulate intermittent reversion from the latent to the infectious state. HSV latency associated transcript (LAT) plays a critical role in latency-reactivation and is the only viral transcript detected at high levels in infected sensory neurons of humans, mice, and rabbits. Because the viral genome, but no detectable infectious virus, is present in latent HSV infection, it is clear that the viral genome is not fully quiescent in latently infected cells. Our previous studies, and those published in the current funding period, established an important link between gD expression and the ability of LAT to enhance herpes virus entry mediator (HVEM) expression in TG of latently infected mice. Our findings suggest a novel model, in which gD expression and LAT enhanced upregulation of HVEM expression contribute to HSV reactivation in TG of latently infected mice. In this proposal, we hypothesize that low level gD expression and its binding to HVEM contribute to ocular HSV reactivation. Our progress during the current funding period has provided novel insights into potential mechanisms by which LAT regulation of HVEM expression defines the extent of latency-reactivation and eye disease. Based on our data, we have formulated a bimodal model in which the increased reactivation effects of sncRNA1&2 during latency are mediated by two different, but inter-related, mechanisms: (i) sncRNA1&2 binding to the HVEM promoter leads to higher levels of HVEM expression; and (ii) higher expression of HVEM enhances gD binding to HVEM and increases reactivation. We will test our model using clinically relevant recombinant viruses in mouse models that control for the complexity of the latent microenvironment in TG and its potential relationships with the primary infection as follows: (1) Determine whether sncRNA1&2 are both required to upregulate HVEM and increase reactivation from latency; and (2) Determine whether gD expression during latency and its binding to HVEM is required for efficient reactivation from latency in TG of latently infected mice. Validation of this hypothetical model will identify previously undescribed mechanisms that contribute to HSV-1 reactivation and will provide the framework for identification of molecular targets and viral immune evasion response that could be exploited to better manage latent HSV infection. CLINICAL SIGNIFICANCE AND