Cellular RNA-binding Zinc Finger Proteins in Viral Infection: Understanding the Rules of Engagement

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Project Summary Viruses such as human immunodeficiency virus-1 (HIV-1) and severe acute respiratory syndrome coronavirus-2 (SARS-CoV2) are the causative pathogens of on-going pandemics. Understanding host-virus interactions is essential to combatting these viruses. Proteomic approaches have identified host proteins that target viral proteins; however, we lack information regarding host factors that target viral RNA (ZAP) or regulate host RNAs (TTP, MCPIP1, and Roquin-1) during infection. Interestingly, the short list of known RNA-binding effectors shares a common CCCH-type zinc finger (ZnF) motif. Unlike other cellular ZnF proteins, most CCCH-type ZnF proteins bind RNA rather than DNA and account for nearly 40% of all RNA-binding ZnF proteins. Furthermore, many of these proteins have poorly annotated functions. I propose that unidentified CCCH ZnF proteins regulate host and viral RNAs during infection. Through systematic siRNA screening, I will identify CCCH-ZnF proteins that modulate the replication of two clinically relevant viruses: human immunodeficiency virus type 1 (HIV-1) and human coronavirus OC43 (HCoV-OC43). I have already identified exciting CCCH-ZnF proteins that have significant beneficial or deleterious effects on HIV-1 and HCoV-OC43 replication. While unique hits were identified for each virus, seven CCCH-ZnF proteins are shared hits in both HIV-1 and HCoV-OC43 screens, suggesting that some of these factors may be broadly antiviral. The proposed research seeks to understand the requirement for RNA-binding by ZnF motif(s) in these host-virus interactions and will use transcriptomics, proteomics, and microscopy approaches to elucidate detailed mechanisms of action. Preliminary results justify this rapid and robust screening method for identification of CCCH-ZnF (and other ZnF-type) factors involved in replication of diverse virus families. Furthermore, I will establish an efficient, streamlined workflow to determine the most promising hits for mechanistic exploration. My work will uncover novel host factors networks that are involved in virus replication and assign molecular functions to cellular CCCH ZnF proteins.