Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Interactions of aCPs with Cytosine-rich Polypyrimidine Tracts Enhance Splicing of Cassette Exons

ABSTRACT: Alternative splicing comprises a robust generator of mammalian transcriptome complexity. Splice site specification and activity are controlled by interactions of cis-acting determinants on a transcript with specific RNA binding proteins. A major subset of these interactions comprises interactions localized to the intronic U-rich polypyrimidine tract located immediately 5â to the majority of splice acceptors. alphaCPs (also referred to as polyC-binding proteins (PCBPs) and hnRNP Es) comprise a subset of KH-domain proteins with high specificity and affinity for C-rich polypyrimidine motifs. Prior studies have revealed that binding of alphaCPs to C-rich motifs can modulate splicing and 3â processing of the human alpha-globin mRNA transcript in the nucleus as well as stabilization of the halpha-globin mRNA in the cytoplasm. In the current report, we demonstrate that alphaCPs have a positive impact on the activity of splice acceptor sites in a defined subset of mammalian transcripts via binding to polypyrimidine tracts that are predominantly C-rich. These findings lead us to conclude that the alphaCPs play a global role in determining the splicing activity and levels of cassette exon inclusion within the mammalian transcriptome. To test the impact of aCP proteins on alternative splicing, aCP proteins were knockdown from K562 cells by siRNA. Since aCP1 and aCP2 have redundent function, we therefore designed siRNAs capable of knockdown both isoform at the same time. 3 aCP1/2 combined knockdown and 3 control siRNA knockdown were performed in K562 cells. RNA-seq were then performed to identify alternative splicing pattern mediated by aCP proteins.

ORGANISM(S): Homo sapiens

SUBMITTER: Yu-Ting Tseng

PROVIDER: E-GEOD-71569 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

αCP binding to a cytosine-rich subset of polypyrimidine tracts drives a novel pathway of cassette exon splicing in the mammalian transcriptome.

Ji Xinjun X Park Juw Won JW Bahrami-Samani Emad E Lin Lan L Duncan-Lewis Christopher C Pherribo Gordon G Xing Yi Y Liebhaber Stephen A SA

Nucleic acids research 20160220 5

Alternative splicing (AS) is a robust generator of mammalian transcriptome complexity. Splice site specification is controlled by interactions of cis-acting determinants on a transcript with specific RNA binding proteins. These interactions are frequently localized to the intronic U-rich polypyrimidine tracts (PPT) located 5' to the majority of splice acceptor junctions. αCPs (also referred to as polyC-binding proteins (PCBPs) and hnRNPEs) comprise a subset of KH-domain proteins with high affini ...[more]

PMID: 26896798

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Project description:Alternative splicing (AS) generates transcript variants by the definition of different exonic and intronic regions and causes a massive expansion of transcriptome diversity. Changes in AS patterns have been found to be linked to manifold biological processes, yet fundamental aspects such as the regulation of AS and the functional implications of altered AS programs largely remain to be addressed. In this work, widespread AS regulation by Arabidopsis Polypyrimidine tract-binding protein homologues (AtPTBs) was revealed. In total 452 AS events derived from 308 distinct genes were found to be responsive to the levels of the splicing factors AtPTB1 and AtPTB2, which predominantly triggered splicing of regulated introns, inclusion of cassette exons, and usage of upstream 5' splice sites. In contrast, alternative 3' splice site events were strongly underrepresented among the AtPTB1/2 targets and no major AS regulatory function of the distantly related AtPTB3 was found. Dependent on their position within the mRNA, AtPTB-regulated events can both modify the untranslated regions and give rise to alternative protein products. Gene ontology analysis revealed a connection of AtPTB-mediated AS control with diverse biological processes, and the functional implications of selected AS events were further elucidated in the context of seed germination and flowering time control. Specifically, AtPTB misexpression changes AS of the PHYTOCHROME INTERACTING FACTOR 6 (PIF6) pre-mRNA, coinciding with altered rates of abscisic acid-dependent seed germination. Furthermore, AS patterns as well as the expression of key flowering regulators were massively changed in an AtPTB1/2 level-dependent manner. In conclusion, our work has revealed widespread AS regulatory functions of the AtPTB splicing factors with important functional implications in various fundamental processes of Arabidopsis development. Analysis of alternative splicing patterns in plants with increased and decreased levels of the 3 Arabidopsis Polypyrimidine-tract binding protein homologues in comparison to wild type samples, determined in duplicates

Dataset Information

Interactions of aCPs with Cytosine-rich Polypyrimidine Tracts Enhance Splicing of Cassette Exons

Publications

αCP binding to a cytosine-rich subset of polypyrimidine tracts drives a novel pathway of cassette exon splicing in the mammalian transcriptome.

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