TRIM25 Binds RNA to Modulate Cellular Anti-viral Defense |
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Authors: | Jacint G Sanchez Konstantin MJ Sparrer Cindy Chiang Rebecca A Reis Jessica J Chiang Matthew A Zurenski Yueping Wan Michaela U Gack Owen Pornillos |
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Institution: | 1. Department of Molecular Physiology and Biological Physics, University of Virginia, Charlottesville, VA, USA;2. Department of Microbiology, University of Chicago, Chicago, IL, USA;3. Department of Microbiology and Immunobiology, Harvard Medical School, Boston, MA, USA |
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Abstract: | TRIM25 is a multi-domain, RING-type E3 ubiquitin ligase of the tripartite motif family that has important roles in multiple RNA-dependent processes. In particular, TRIM25 functions as an effector of RIG-I and ZAP, which are innate immune sensors that recognize viral RNA and induce ubiquitin-dependent anti-viral response mechanisms. TRIM25 is reported to also bind RNA, but the molecular details of this interaction or its relevance to anti-viral defense have not been elucidated. Here, we characterize the RNA-binding activity of TRIM25 and find that the protein binds both single-stranded and double-stranded RNA. Multiple regions of TRIM25 contribute to this functionality, including the C-terminal SPRY domain and a lysine-rich motif in the linker segment connecting the SPRY and coiled-coil domains. RNA binding modulates TRIM25's ubiquitination activity in vitro, its localization in cells, and its anti-viral activity. Taken together with other studies, our results indicate that RNA binding by TRIM25 has at least three important functional consequences: by enhancing ubiquitination activity, either through allosteric effects or through clustering of multiple TRIM25 molecules; by modulating the multi-domain structure of the TRIM25 dimer, and thereby structural coupling of the SPRY and RBCC elements during the ubiquitination reaction; and by facilitating subcellular localization of the E3 ligase during virus infection. |
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Keywords: | ubiquitination RNA recognition signaling IFN interferon PEI polyethyleneimine WT wild-type SEC-MALS size exclusion chromatography coupled with multi-angle light scattering EMSA electrophoretic mobility shift assay KO knockout DENV dengue virus VSV-eGFP eGFP-expressing vesicular stomatitis virus IAV influenza A virus NS1 non-structural protein 1 WCL whole cell lysate GST-2CARD GST-fused RIG-I 2CARD PBS phosphate-buffered saline |
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