ABSTRACT Kaposi sarcoma-associated herpesvirus (KSHV) is the etiologic agent of the majority of AIDS- associated cancers. It is endemic in many areas of Africa where, due to the extraordinarily high HIV burden, Kaposi sarcoma has emerged as one of the most common cancers on the continent. During AIDS- induced immunosuppression, KSHV replication is no longer effectively controlled, and, together with a large latently infected population of cells, contributes to disease progression. During lytic replication, KSHV and the closely related model murine gammaherpesvirus MHV68 dramatically remodel the host gene expression environment. Key to this remodeling is its virally encoded, messenger RNA (mRNA) specific endonuclease termed SOX, which accelerates degradation of a broad spectrum of mRNAs. SOX activity plays diverse roles in the in vivo gammaherpesvirus lifecycle and immune evasion. However, the mechanisms underlying the RNA target specificity of SOX remain largely unknown, despite their prominent roles in shaping the mRNA abundance profile during infection. Our data show that SOX uses a combination of RNA sequence and structure to capture a broad set of mRNA targets while preserving selectivity. In Aim 1, we will probe how novel protein-protein interactions between SOX and components of the RNA processing machinery influence the susceptibility of mRNAs to endonuclease targeting across a range of cell types. We will then define the downstream consequences of mRNA targeting by SOX, including how large scale changes to mRNA degradation cause profound ‘ripple effects’ to the gene expression landscape. In Aim 2, we will mechanistically characterize a new pathway we discovered in mammalian cells that functionally links the last stage of the mRNA lifecycle (degradation) to the first stage (transcription). This mRNA decay-transcription “feedback” pathway is activated by SOX and results in a large-scale reduction of RNA polymerase II occupancy selectively across the mammalian but not the viral genome. Findings derived from this proposal should have a sustained impact on the field of gammaherpesvirus biology, and change current perceptions on how stress or virus-induced alterations to mRNA stability influence seemingly distal components of the gene regulation circuitry.