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PsiCLIP of SmB, Prp16 and Prp22 in C, C* and P spliceosomal complexes

ABSTRACT: Despite the major progress made into spliceosome mechanisms through CryoEM, a major obstacle of this approach is its inability to map the positioning of helicases on the RNA substrate. Here we present a method ‘psiCLIP’ which probes protein-RNA interactions in specific spliceosomal states. The method is based on iCLIP, but is different to previous iCLIP studies in that it is performed on step-specific spliceosomes that are prepared from in vitro splicing extracts, similarly to those prepared for CryoEM. We applied psiCLIP to SmB (C complex), Prp16 (C complex) and Prp22 (C* and P complexes), using pre-mRNA substrates based on UBC4 and ACT1. We also used ATPase deficient dominant negative (dn) mutants of Prp16 and Prp22 to determine the contribution of ATPase activity to binding patterns.

INSTRUMENT(S): Illumina HiSeq 2500

ORGANISM(S): Saccharomyces cerevisiae

SUBMITTER: Charlotte Capitanchik

PROVIDER: E-MTAB-8895 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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