RNA Sequence Context Effects Measured In Vitro Predict In Vivo Protein Binding and Regulation.
Many RNA binding proteins (RBPs) bind specific RNA sequence motifs, but only a small fraction (∼15%-40%) of RBP motif occurrences are occupied in vivo. To determine which contextual features discriminate between bound and unbound motifs, we performed an in vitro binding assay using 12,000 mouse RNA sequences with the RBPs MBNL1 and RBFOX2. Surprisingly, the strength of binding to motif occurrences in vitro was significantly correlated with in vivo binding, developmental regulation, and evolutionary age of alternative splicing. Multiple lines of evidence indicate that the primary context effect that affects binding in vitro and in vivo is RNA secondary structure. Large-scale combinatorial mutagenesis of unfavorable sequence contexts revealed a consistent pattern whereby mutations that increased motif accessibility improved protein binding and regulatory activity. Our results indicate widespread inhibition of motif binding by local RNA secondary structure and suggest that mutations that alter sequence context commonly affect RBP binding and regulation.
['Taliaferro|J Matthew|JM|', 'Lambert|Nicole J|NJ|', 'Sudmant|Peter H|PH|', 'Dominguez|Daniel|D|', 'Merkin|Jason J|JJ|', 'Alexis|Maria S|MS|', 'Bazile|Cassandra|C|', 'Burge|Christopher B|CB|']
D000465:Algorithms D017398:Alternative Splicing D000818:Animals D001665:Binding Sites D002417:Cattle D002454:Cell Differentiation D004268:DNA-Binding Proteins D005347:Fibroblasts D015870:Gene Expression D008251:Macaca D051379:Mice D018345:Mice, Knockout D000066450:Mouse Embryonic Stem Cells D009154:Mutation D009474:Neurons D009690:Nucleic Acid Conformation D059372:Nucleotide Motifs D011485:Protein Binding D054730:Protein Interaction Domains and Motifs D012313:RNA D000072260:RNA Splicing Factors D016601:RNA-Binding Proteins D051381:Rats D012984:Software
RBNS RNA processing post-transcriptional