Circular RNAs in HSV1 infection and antiviral immunity

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Project Summary Circular (circ) RNAs are a recently described class of RNAs generated by backsplicing of adjacent exons, whose expression is altered in several disease states and some of the few viral infections in which they have been studied. Functionally, circRNAs rarely direct protein synthesis but have been linked to the sequestration of microRNAs and RNA binding proteins (RNA-BPs). CircRNAs were recently implicated in controlling the activation of double-stranded (ds) RNA-activated eIF2α kinase PKR by providing an abundant pool of inhibitory short duplex RNA sequences. Bound inactive PKR can be rapidly mobilized by if RNase L is activated, resulting in endoribonucleolytic cleavage of circRNAs. Generated by both RNA virus replication and dsDNA virus transcription from opposing genome strands, dsRNA is a potent pathogen associated molecular pattern (PAMP) whose recognition by PKR and the OAS/RNase L pathway leads to an abrupt defensive shutdown of mRNA translation to thwart viral protein synthesis. In addition to possible virus and circRNA -specific interactions, the potential of circRNAs to calibrate dsRNA responses therefore suggests a broad significance to virus biology. However, precisely how the infected cell circRNA population responds to herpes simplex type 1 (HSV1) infection and impacts virus reproduction remains unknown and represents a significant knowledge gap. The overall goal of this proposal is to understand how circRNAs contribute to HSV1 infection and antiviral immunity. Our preliminary data in HSV1-infected cells indicate that circRNAs are protected from degradation by the viral endoribonuclease vhs and are in fact selectively upregulated through a mechanism dependent on the viral immediate early protein ICP27. We also find that PKR activation is dependent on RNase L function, suggesting that circRNAs are important inhibitors of PKR in HSV1 infection that must be degraded for a full antiviral response. These data are consistent with the novel hypothesis that HSV1 actively manipulates this class of host RNAs to evade cell intrinsic antiviral defenses and preserve protein synthesis and viral replication. This proposal will be divided into two aims to fully understand 1) how HSV1 shapes the infected cell circRNAome and 2) the functional impact of these alterations. These studies will reveal how circRNAs are manipulated by HSV1 and their importance in infection and immunity to a medically important human pathogen. Ultimately, this work could lead to new strategies for treating a wide variety of viral infections and have significant implications for our understanding of infection, host defenses and circRNA biology.