Restriction of KSHV by cellular RNA decay pathways

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Project Summary Kaposi’s sarcoma-associated herpesvirus (KSHV) is a member of the subfamily gammaherpesvirinae and is causally associated with the development of several malignancies including Kaposi’s sarcoma and primary effusion lymphoma (PEL). The virus’ ability to establish latency as well as reactivate are essential for the development of KSHV-associated disease. Despite these requirements for disease progression there are significant gaps in knowledge regarding cell-intrinsic mechanisms that restrict the KSHV lifecycle. While innate restriction is typically thought of in the context of antiviral immune responses, growing evidence suggests that cellular RNA quality control pathways have an antiviral role. Nonsense-mediated RNA decay (NMD) is an evolutionarily conserved RNA decay pathway that facilitates degradation of RNAs on which ribosomes are deemed to terminate translation aberrantly. Emerging evidence has pointed to a role for NMD in antiviral restriction with a prominent role limiting replication of positive-stranded RNA viruses, however, a role for NMD in DNA virus restriction, such as KSHV, was unknown. We recently reported the discovery that NMD is a cell- intrinsic restriction mechanism for DNA viruses and demonstrated that it imposes a significant restriction on the KSHV lifecycle in PEL cells. Our data demonstrate that NMD-dependent restriction is linked both to the regulation of the unfolded protein response (UPR) as well as targeted degradation of the main KSHV transcription factor, RTA. Building upon these observations our central hypothesis is that NMD restricts KSHV reactivation by targeting key viral mRNAs as well as cellular transcripts in pathways important for viral gene expression and that the virus antagonizes NMD through viral-encoded mechanisms. To test this hypothesis, we propose an integrated series of experiments aimed at determining the interactions between NMD and KSHV. In Aim 1, we will investigate how NMD-dependent regulation of the UPR pathway regulates the KSHV lifecycle. In Aim 2, we will investigate a class of KSHV mRNAs that escape NMD despite harboring sequence features that should render them NMD-susceptible. In Aim 3, we will determine the mechanism by which KSHV-encoded proteins inhibit NMD. Completion of these studies is expected to determine how NMD restricts the KSHV lifecycle as well as the mechanisms the virus employs to antagonize it. These will represent fundamental new insights into how DNA virus infection is regulated by cell-intrinsic mechanisms and can be harnessed for the development of new therapeutic strategies.