Journal
BIOCHIMICA ET BIOPHYSICA ACTA-GENE REGULATORY MECHANISMS
Volume 1839, Issue 1, Pages 25-32Publisher
ELSEVIER SCIENCE BV
DOI: 10.1016/j.bbagrm.2013.11.006
Keywords
Caspase-2; SRSF3; Apoptotic; Anti-apoptotic; Pre-mRNA splicing; Exon 9
Categories
Funding
- Mid-career Researcher Program through a National Research Foundation (NRF) grant [2011-0016757]
- Ministry of Education, Science, and Technology (MEST), Korea
- Gwangju Institute of Science and Technology (GIST)
- Ministry of Science, ICT & Future Planning, Republic of Korea [GIST-08] Funding Source: Korea Institute of Science & Technology Information (KISTI), National Science & Technology Information Service (NTIS)
- National Research Foundation of Korea [21A20131212125, 2011-0016757] Funding Source: Korea Institute of Science & Technology Information (KISTI), National Science & Technology Information Service (NTIS)
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Alternative splicing plays an important role in gene expression by producing different proteins from a gene. Caspase-2 pre-mRNA produces anti-apoptotic Casp-2S and pro-apoptotic Casp-2L proteins through exon 9 inclusion or skipping. However, the molecular mechanisms of exon 9 splicing are not well understood. Here we show that knockdown of SRSF3 (also known as SRp20) with siRNA induced significant increase of endogenous exon 9 inclusion. In addition, overexpression of SRSF3 promoted exon 9 skipping. Thus we conclude that SRSF3 promotes exon 9 skipping. In order to understand the functional target of SRSF3 on caspase-2 pre-mRNA, we performed substitution and deletion mutagenesis on the potential SRSF3 binding sites that were predicted from previous reports. We demonstrate that substitution mutagenesis of the potential SRSF3 binding site on exon 8 severely disrupted the effects of SRSF3 on exon 9 skipping. Furthermore, with the approach of RNA pulldown and immunoblotting analysis we show that SRSF3 interacts with the potential SRSF3 binding RNA sequence on exon 8 but not with the mutant RNA sequence. In addition, we show that a deletion of 26 nt RNA from 5 ' end of exon 8, a 33 nt RNA from 3 ' end of exon 10 and a 2225 nt RNA from intron 9 did not compromise the function of SRSF3 on exon 9 splicing. Therefore we conclude that SRSF3 promotes exon 9 skipping of caspase-2 pre-mRNA by interacting with exon 8. Our results reveal a novel mechanism of caspase-2 pre-mRNA splicing. (C) 2013 Elsevier B.V. All rights reserved.
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