iCLIP reveals the function of hnRNP particles in splicing at individual nucleotide resolution
Julian König,Kathi Zarnack,Gregor Rot,Tomaž Curk,Melis Kayikci,Blaž Zupan,Daniel J. Turner,Nicholas M. Luscombe,Jernej Ule +8 more
TLDR
Individual-nucleotide resolution UV cross-linking and immunoprecipitation (iCLIP) data show that hnRNP C recognizes uridine tracts with a defined long-range spacing consistent with hmRNP particle organization, and integration of transcriptome-wide iCLIP data and alternative splicing profiles into an 'RNA map' indicates how the positioning of hn RNP particles determines their effect on the inclusion of alternative exons.Abstract:
In the nucleus of eukaryotic cells, nascent transcripts are associated with heterogeneous nuclear ribonucleoprotein (hnRNP) particles that are nucleated by hnRNP C. Despite their abundance, however, it remained unclear whether these particles control pre-mRNA processing. Here, we developed individual-nucleotide resolution UV cross-linking and immunoprecipitation (iCLIP) to study the role of hnRNP C in splicing regulation. iCLIP data show that hnRNP C recognizes uridine tracts with a defined long-range spacing consistent with hnRNP particle organization. hnRNP particles assemble on both introns and exons but remain generally excluded from splice sites. Integration of transcriptome-wide iCLIP data and alternative splicing profiles into an 'RNA map' indicates how the positioning of hnRNP particles determines their effect on the inclusion of alternative exons. The ability of high-resolution iCLIP data to provide insights into the mechanism of this regulation holds promise for studies of other higher-order ribonucleoprotein complexes.read more
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Donny D. Licatalosi,Aldo Mele,John J. Fak,Jernej Ule,Melis Kayikci,Sung Wook Chi,Tyson A. Clark,Anthony C. Schweitzer,John E. Blume,Xuning Wang,Jennifer C. Darnell,Robert B. Darnell +11 more
TL;DR: A genome-wide means of mapping protein–RNA binding sites in vivo, by high-throughput sequencing of RNA isolated by crosslinking immunoprecipitation (HITS-CLIP), which revealed a large number of Nova–RNA interactions in 3′ untranslated regions, leading to the discovery that Nova regulates alternative polyadenylation in the brain.