Poster abstracts

Poster number 11 submitted by Samarth Kumar

The function of specific MAVS-associated RNAs in antiviral signaling

Samarth Kumar (Division of Immunobiology, CCHMC, University of Cincinnati), Juan R. Molina (Molecular Genetics, University of Cincinnati), Andrew Herr (Division of Immunobiology, CCHMC, University of Cincinnati), Nandan Gokhale (Division of Immunobiology, CCHMC, University of Cincinnati)

Abstract:
MAVS, the adaptor protein for antiviral RIG-I-like receptors (RLRs), forms a higher-order signaling complex that mediates antiviral signaling by inducing expression of interferons (IFNs) to restrict viruses. MAVS signaling is crucial for protection against viruses but must be finely tuned as excessive activation can lead to inflammation and autoimmunity. Our lab has identified cellular RNA interaction as a novel regulatory layer that promotes MAVS signaling. However, the precise biochemical mechanisms underlying the control of MAVS signaling by RNA remain a gap in knowledge. MAVS interacts with the 3′ untranslated regions (UTRs) of cellular mRNAs through a central intrinsically disordered region (IDR). IDRs interactions with nucleic acids can drive or stabilize liquid-liquid phase separated (LLPS) condensates. MAVS can undergo LLPS, but whether its IDR and RNA-binding activity contribute this process is unknown. I hypothesize that the interaction of specific MAVS-binding 3′ UTRs like PMAIP1 and IFIT2 with the MAVS IDR promotes MAVS phase separation to augment antiviral signaling. Towards testing this, I have used irCLIP and sucrose gradient centrifugation to determine that positively charged residues in the MAVS IDR mediate RNA interaction. Mutating these residues diminishes RLR signaling. We now aim to purify this RNA-binding mutant MAVS and WT MAVS from bacteria and analyze MAVS phase separation in the presence of specific 3′ UTRs and test the ability of MAVS-RNA condensates to promote MAVS signaling in an in vitro assay. To further determine the specificity of 3′ UTR interactions with MAVS, I aim to test the impact of tethering these or control UTRs to MAVS using the MS2-MCP RNA stem loop system as well as UTR deletion and complementation strategies on MAVS signaling.

Keywords: RNA, MAVS, 3 UTR