7. PROJECT SUMMARY / ABSTRACT Regulation of gene expression at the level of translation allows cells and organisms to respond swiftly to physiological stress and changing environments. Indeed, differential translation of capped, polyadenylated mRNAs by eukaryotic ribosomes plays a critical role in biological processes vital for human health, including normal cell growth, differentiation, development, learning and memory, and responses to physiological stress including virus infection. Viral model systems have proven useful in elaborating cellular translational control strategies because their replication is absolutely reliant upon virus mRNA translation by host ribosomes. Here, we utilize a herpesvirus family member, human cytomegalovirus (HCMV), to probe the complex circuitry regulating mRNA translation. Although innocuous in healthy individuals, HCMV is a widespread, opportunistic pathogen responsible for severe disease among the immunocompromised, including organ transplant recipients and AIDS patients. In addition, congenital HCMV infection is the leading viral cause of birth defects in newborns. Our long-term overall objective is to understand the mechanism(s) through which HCMV manipulates the cellular translational machinery to control viral replication. A fundamental step regulating protein synthesis in HCMV-infected cells involves ribosome recruitment to the 5'-end of m7G-capped virus and host mRNAs. This typically involves 5'-cap-recognition by a translation initiation factor complex containing the cap-binding protein eIF4E followed by loading the eIF3-bound 40S ribosome. Unexpectedly, an alternative form of 40S ribosome loading relying upon cap-recognition by a specific eIF3 subunit, eIF3d, has been described that bypasses the need for eIF4E and is distinctly regulated. Preliminary results establish that targeting eIF3d selectively inhibited HCMV replication, reduced polyribosome abundance and interfered with expression of essential virus late genes and a host gene expression signature indicative of chronic ER stress that fosters HCMV reproduction. Based on our preliminary results, we hypothesize that eIF3d-dependent cap- recognition is hijacked to exploit virus-induced ER-stress and switching between eIF4E and eIF3d-responsive translation differentially tunes virus and host gene expression in infected cells. This hypothesis is tested in three specific aims designed to: i) determine how HCMV infection impacts eIF3d accumulation and sub-cellular distribution; ii) define the mechanism whereby eIF3d selectively shapes the gene expression landscape in HCMV-infected cells; and iii) decipher how eIF3d activity is regulated in HCMV-infected cells. The project is impactful because how non-canonical translation initiation mechanisms involving eIF3d regulate differential mRNA translation during stress responses like virus infection remains largely unknown and represents a significant knowledge gap poised to reveal new biology and therapeutic opp...