The RNA-binding profile of Acinus, a peripheral component of the exon junction complex, reveals its role in splicing regulation.

Julie Rodor,Benjamin J Blencowe,Eduardo Eyras,Qun Pan,Javier F Cáceres

doi:10.1261/rna.057158.116

Abstract

Acinus (apoptotic chromatin condensation inducer in the nucleus) is an RNA-binding protein (RBP) originally identified for its role in apoptosis. It was later found to be an auxiliary component of the exon junction complex (EJC), which is deposited at exon junctions as a consequence of pre-mRNA splicing. To uncover the cellular functions of Acinus and investigate its role in splicing, we mapped its endogenous RNA targets using the cross-linking immunoprecipitation protocol (iCLIP). We observed that Acinus binds to pre-mRNAs, associating specifically to a subset of suboptimal introns, but also to spliced mRNAs. We also confirmed the presence of Acinus as a peripheral factor of the EJC. RNA-seq was used to investigate changes in gene expression and alternative splicing following siRNA-mediated depletion of Acinus in HeLa cells. This analysis revealed that Acinus is preferentially required for the inclusion of specific alternative cassette exons and also controls the faithful splicing of a subset of introns. Moreover, a large number of splicing changes can be related to Acinus binding, suggesting a direct role of Acinus in exon and intron definition. In particular, Acinus regulates the splicing of DFFA/ICAD transcript, a major regulator of DNA fragmentation. Globally, the genome-wide identification of RNA targets of Acinus revealed its role in splicing regulation as well as its involvement in other cellular pathways, including cell cycle progression. Altogether, this study uncovers new cellular functions of an RBP transiently associated with the EJC.

Highlights

There is an extensive coupling among different steps in eukaryotic gene expression, as shown by the intimate connection between transcription and RNA processing (Braunschweig et al 2013; Bentley 2014)
Consistent with Acinus harboring an RNA recognition motif (RRM), it was recently shown that it crosslinked to RNA as part of the mRNA–protein interactome of HeLa cells (Castello et al 2012)
In order to uncover the roles of Acinus in RNA processing, we decided to focus on the identification of its endogenous RNA targets

Summary

Introduction

There is an extensive coupling among different steps in eukaryotic gene expression, as shown by the intimate connection between transcription and RNA processing (Braunschweig et al 2013; Bentley 2014). RBPs have a profound impact on cellular gene expression networks, affecting processes as diverse as transcription, constitutive and alternative pre-mRNA splicing, microRNA (miRNA) biogenesis and function and mRNA translation. New protein components of messenger RNA ribonucleoparticles (mRNPs) have been recently identified (Baltz et al 2012; Castello et al 2012; Mitchell and Parker 2014). Some of these RBPs associate with the mRNA in a splicingdependent manner (Merz et al 2007).

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Results

Conclusion