The capacity of eukaryotic cells to respond to numerous forms of stress, ranging from thermal intolerance or nutrient deprivation to viral infection, involves, in part, their ability to regulate the activity of elF2, a critical translation initiation factor. Upon phosphorylation on it's alpha subunit by one of four known elF2alpha kinases, elF2 is inactivated and translation is inhibited. The overall long - term objective of this project is to understand how elF2alpha phosphorylation is regulated in response to stress generated by viral infection. In infected cells, the appropriation and redirection of many cellular functions coupled with the rapid accumulation of viral gene products induces cellular stress and activates host defenses. Copious quantities of double stranded (ds)RNA produced in virus infected cells activates the cellular elF2alpha kinase PKR, while the synthesis of viral glycoproteins increases the load on the endoplasmic reticulum, exceeding its capacity to correctly fold and process ER client proteins. This latter condition triggers the unfolded protein response (UPR) and the ensuing phosphorylation of elF2alpha arrests translation. In this proposal, we investigate viral strategies that prevent host defenses from inactivating the translation factor elF2. Our studies focus on herpes simplex virus-1, a neurotrophic herpesvirus whose productive replication is responsible for a spectrum of diseases, ranging from epithelial sores, severe ocular disease and life threatening encephalitis in immunocompetent hosts to disseminated disease in neonates and immunocompromised individuals. Both the gamma(1)34.5 and Us11 gene products are known to regulate elF2alpha phosphorylation. While the gamma(1)34.5 gene product recruits a cellular phosphatase to dephosphorylate elF2alpha, Us11 prevents activation of the cellular elF2alpha kinase PKR via an unknown mechanism. Furthermore, HSV-1 expresses a previously uncharacterized function, distinct from the polypeptide products of the Us11and gamma(1)34.5 genes, that confers resistance to ER stress. We will (i) evaluate the effects of HSV-1 infection on ER stress transducers; (ii) identify the gene product (s) responsible for preventing elF2alpha phosphorylation in response to effectors that stimulate the UPR; and (iii) investigate the molecular mechanisms by which Us11 prevents PKR activation.
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