Investigation of the role of HDAC activity in regulation of HCMV replication in the salivary epithelium

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The human cytomegalovirus (HCMV) is a significant public health concern in the United States. Most important are the effects of the virus on developing fetuses and immunocompromised individuals where it causes a variety of pathological conditions ranging in severity from mild to life-threatening. Since HCMV is present in a persistent or latent form in 50-90% of the world’s adult population, the identification of viral gene products and mechanisms that contribute to viral trafficking, persistence, and horizontal transmission is an intense and important area of investigation. We have recently generated and published an in vitro model for studying HCMV replication in primary salivary derived epithelial cells called “salispheres”. These salisphere cells express genes typical of salivary acinar epithelial cells, mimic the physiology of the salivary gland, and serve as a valuable model to understand the basic parameters and mechanisms underlying HCMV replication in the salivary epithelium. Our exciting preliminary data indicates that HCMV strains deficient for the pentamer glycoprotein complex can be rescued for infection of salivary cells using histone deacetylase (HDAC) inhibitors. Moreover, our preliminary data suggest that the pentamer may be required to induce nuclear mobilization of pp71, where it can interact with the HDACs, inhibit their activity, and facilitate lytic replication. We also present preliminary data indicating the viral encoded GPCRs (vGPCRs) are essential for replication in salivary cells and that pharmacological HDAC inhibition can similarly rescue the defect exhibited by vGPCR null viruses. Our data suggest novel roles for both HCMV pentamer and vGPCRs in facilitating lytic replication in salivary cells. Based on our preliminary data, we hypothesize that the HCMV pentamer, vGPCRs, and pp71 work in concert to facilitate efficient viral replication in salivary epithelial cells leading to amplification and spread of virus from the salivary gland into the saliva. The proposed studies are highly significant to cytomegalovirus pathogenesis as the salivary gland and its secretions play an important role in horizontal transmission of virus, yet little is known about the viral mechanisms that facilitate infection and replication within the salivary epithelium. In aim 1, we will test whether the HCMV pentamer complex induces mobilization of the pp71 tegument protein into the nucleus to facilitate HDAC inhibition. In aim 2, we will test whether knockdown of HDAC1 or HDAC3 expression or overexpression of pp71 can rescue pentamer null viruses for efficient replication in salivary cells. In aim 3, we will determine whether HCMV vGPCR signaling works in concert with pentamer and pp71 to drive HCMV lytic replication in the salivary epithelial cells. The innovative experiments proposed in this application will generate important insight into the molecular and physiological properties of HCMV involved in salivary epithelial cell infection. Defining the mechanisms underlying salivary gland replication and spread could ultimately lead to the development of unique antivirals designed to prevent cytomegalovirus transmission via saliva.