Abstract
The PolyC binding proteins (PCBPs) impact alternative splicing of a subset of mammalian genes that are enriched in basic cellular functions. Here, we focus our analysis on PCBP-controlled cassette exon-splicing within the cell cycle control regulator cyclin-dependent kinase-2 (CDK2) transcript. We demonstrate that PCBP binding to a C-rich polypyrimidine tract (PPT) preceding exon 5 of the CDK2 transcript enhances cassette exon inclusion. This splice enhancement is U2AF65-independent and predominantly reflects actions of the PCBP1 isoform. Remarkably, PCBPs’ control of CDK2 ex5 splicing has evolved subsequent to mammalian divergence via conversion of constitutive exon 5 inclusion in the mouse CDK2 transcript to PCBP-responsive exon 5 alternative splicing in humans. Importantly, exclusion of exon 5 from the hCDK2 transcript dramatically represses the expression of CDK2 protein with a corresponding perturbation in cell cycle kinetics. These data highlight a recently evolved post-transcriptional pathway in primate species with the potential to modulate cell cycle control.
Highlights
RNA splicing is a highly dynamic process that is regulated by a broad array of RNA binding proteins (RBP)
We find that splicing of cyclindependent kinase-2 (CDK2) exon 5 is positively regulated by PolyC binding proteins (PCBPs) and that this control is mediated via interactions of PCBPs with a C-rich polypyrimidine tract (PPT) splice acceptor 5 to exon 5
These studies further revealed that exons enhanced by PCBPs are preceded at a high frequency by PPTs that are markedly enriched for C residues
Summary
RNA splicing is a highly dynamic process that is regulated by a broad array of RNA binding proteins (RBP) Many of these RBPs constitute core components of mammalian spliceosome complexes [1] or interact with these complexes to modulate assembly of the splicing machinery [2]. The activity of a splice acceptor site located 5 to an exon is a frequent target of splicing control and its activity is often determined by the efficiency with which RNP complexes assemble at the polypyrimidine tract (PPT) located immediately 5 of the AG splice acceptor dinucleotide. Interactions of RNA binding proteins to a PPT can alter initial U2AF65 binding, splicing complex assembly, and/or provide alternative pathways to activate splice acceptor functions [3,6,7,8]
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