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Dissecting noncoding and pathogen RNA-protein interactomes.

RNA (New York, N.Y.) (2014-11-21)
Ryan A Flynn, Lance Martin, Robert C Spitale, Brian T Do, Selena M Sagan, Brian Zarnegar, Kun Qu, Paul A Khavari, Stephen R Quake, Peter Sarnow, Howard Y Chang
要旨

RNA-protein interactions are central to biological regulation. Cross-linking immunoprecipitation (CLIP)-seq is a powerful tool for genome-wide interrogation of RNA-protein interactomes, but current CLIP methods are limited by challenging biochemical steps and fail to detect many classes of noncoding and nonhuman RNAs. Here we present FAST-iCLIP, an integrated pipeline with improved CLIP biochemistry and an automated informatic pipeline for comprehensive analysis across protein coding, noncoding, repetitive, retroviral, and nonhuman transcriptomes. FAST-iCLIP of Poly-C binding protein 2 (PCBP2) showed that PCBP2-bound CU-rich motifs in different topologies to recognize mRNAs and noncoding RNAs with distinct biological functions. FAST-iCLIP of PCBP2 in hepatitis C virus-infected cells enabled a joint analysis of the PCBP2 interactome with host and viral RNAs and their interplay. These results show that FAST-iCLIP can be used to rapidly discover and decipher mechanisms of RNA-protein recognition across the diversity of human and pathogen RNAs.

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Sigma-Aldrich
Anti-PCBP2 antibody produced in rabbit, Prestige Antibodies® Powered by Atlas Antibodies, affinity isolated antibody, buffered aqueous glycerol solution