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

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.

Human nuclear RNAi-defective 2 (NRDE2) is an essential RNA splicing factor

“…2A). ACIN1 is a peripheral EJC component that has been shown to preferentially bind short, GC-rich introns with weak splice signals (Rodor et al 2016)-precisely the types of introns retained following NRDE2 depletion. It will be interesting to determine whether the intron specificity of NRDE2 is intrinsic or mediated by its protein partners.…”

U2 AF 65 assemblies drive sequence‐specific splice site recognition

“…Conserved cassette exon in human and mouse was retained by joining chromosome coordinates after converting mouse coordinates to human. The corresponding exon–exon junctions were then used as a library for Bowtie alignment of published RNA‐Seq sequences for HeLa cells in standard conditions (~ 160 M reads) . Splicing events with at least two reads aligned on both the skipped and inclusion isoforms and with a sum of at least 20 reads aligning on both isoforms were retained.…”

“…Exons were filtered for the absence of alternative 5 0 and 3 0 splice site, for the existence of unique 5 0 splice site for the preceding and 3 0 splice site for the following intron in all annotated transcripts, and finally for the exons being present in at least one and absent in at least one of the annotated transcripts as depicted in Appendix Fig S6. Conserved cassette exon in human and mouse was retained by joining chromosome coordinates after converting mouse coordinates to human. The corresponding exon-exon junctions were then used as a library for Bowtie alignment of published RNA-Seq sequences for HeLa cells in standard conditions (~160 M reads) [69]. Splicing events with at least two reads aligned on both the skipped and inclusion isoforms and with a sum of at least 20 reads aligning on both isoforms were retained.…”

Genome Sequencing and RNA-Motif Analysis Reveal Novel Damaging Noncoding Mutations in Human Tumors

“…We collected binding sites from 91 CLIP-Seq experiments for 68 different RBPs from multiple sources (34)(35)(36)(37)(38)(39)(40) and selected the available significant CLIP clusters from each experiment. For datasets with two replicates, we selected the intersecting regions of the significant CLIP clusters, i.e., the genomic ranges covered by both replicates.…”