Understanding Epstein Barr virus oncogenicity through non-coding RNA structure

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 DESCRIPTION (provided by applicant): Epstein-Barr virus (EBV) is a herpesvirus that causes lifelong latent infections in ~95% of humans. In ways not yet fully known, EBV latency is implicated in cancers such as Burkitt's and Hodgkin's lymphomas. An understudied aspect of EBV biology, that may help us to understand its oncogenic properties, is the role played by RNA structure. My research in the Steitz lab predicts that approximately 30% of the EBV genome generates RNA with thermodynamically stable and evolutionarily conserved RNA structure. This suggests that a substantial fraction of the EBV transcriptome may have unknown functional roles mediated by non-coding (nc)RNA structure. Likewise, structured human host transcripts may also play key roles in EBV infection and disease. This project will collect transcriptome-wide biochemical structure probing data from living human cells: uninfected and infected with EBV. These data will be used to model RNA structure in latent and lytic virus/host transcripts and to identify important RNA structural dynamics with implications for EBV infection and cancer. This project will also explore the possible targeting of RNA structure for therapeutic intervention. In the K99 phase, the transcriptome of latent EBV-infected human cells will be chemically probed using RNA structure-sensitive cell-permeable reagents. Using novel techniques developed in the lab of my co-mentor Prof. Philip Bevilacqua, reactive sites will be read out using RNA-Seq. These data will provide an unprecedented view of RNA structure in EBV latency. The functions of two highly conserved, structured, and abundant latency- associated ncRNAs, which I recently discovered, will also be investigated. These RNAs are produced during a highly oncogenic form of latency (latency III) and this analysis may help explain the cancer-causing properties of latency III. These endeavors lay the groundwork for future studies of virus and host RNA structures important to EBV infection and cancer, which I will make the focus of my future lab. In the R00 phase, the skills and methods I will develop in the K99 phase will be applied to study RNA structure in the transcriptomes of EBV-negative and lytically re-activated EBV-infected cells. In addition to shedding light on RNA structures in the lytic EBV transcriptome, this work will allow comparisons to be made between structures present in EBV-negative, latent, and lytic host and EBV transcriptomes. This will identify potential structural dynamics in viral and host RNAs, which may be important to infection. In addition to providing fundamentally important basic knowledge about EBV, this research also has significant implications to human health. A better understanding of RNA structure in EBV infection may identify oncogenic RNAs that can be targeted with therapeutics. For example, in collaboration with the Matthew Disney lab, precursor miRNA hairpins generating cancer-associated viral miRNAs will be targeted with RNA-binding small molecules to inhibit miRNA maturation. By advancing the use of RNA-targeting small molecules, this work has the potential to generate new therapeutics aimed at treating EBV-related cancers.