Abstract
The exon junction complex (EJC) that is deposited onto spliced mRNAs upstream of exon–exon junctions plays important roles in multiple post-splicing gene expression events, such as mRNA export, surveillance, localization, and translation. However, a direct role for the human EJC in pre-mRNA splicing has not been fully understood. Using HeLa cells, we depleted one of the EJC core components, Y14, and the resulting transcriptome was analyzed by deep sequencing (RNA-Seq) and confirmed by RT–PCR. We found that Y14 is required for efficient and faithful splicing of a group of transcripts that is enriched in short intron-containing genes involved in mitotic cell-cycle progression. Tethering of EJC core components (Y14, eIF4AIII or MAGOH) to a model reporter pre-mRNA harboring a short intron showed that these core components are prerequisites for the splicing activation. Taken together, we conclude that the EJC core assembled on pre-mRNA is critical for efficient and faithful splicing of a specific subset of short introns in mitotic cell cycle-related genes.
Highlights
Pre-mRNA splicing, the correct and precise removal of introns is an essential part of gene expression in eukaryotes
The exon junction complex (EJC) is composed of four core components, eIF4AIII, Y14, MAGOH and MLN51, and many proteins that are weakly associated with the EJC core, termed EJC peripheral factors
To investigate whether the EJC is implicated in pre-mRNA splicing, we performed a deep-sequencing analysis of transcriptome in Y14-knockdown HeLa cells, i.e., resulting transcriptome was analyzed by deep sequencing (RNA-Seq) analysis
Summary
Pre-mRNA splicing, the correct and precise removal of introns is an essential part of gene expression in eukaryotes. It was shown that core and peripheral EJC components regulate alternative splicing of BCL-X pre-mRNA through its binding to a cis-acting element, whose activity is distinct from the established EJC function [15]. All this evidence suggested that the EJC core is capable of recruiting various splicing regulators and that these interactions may regulate pre-mRNA splicing. A tethering assay of the EJC core components (eIF4AIII, Y14 or MAGOH) demonstrated that the formation of the EJC core onto pre-mRNA (not onto mRNA) enhances splicing These results provide a considerable insight into the EJC-mediated splicing fine-tuning mechanism for short introns in functionally related genes
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