title: Exon 9 skipping of apoptotic caspase-2 pre-mRNA is promoted by SRSF3 through interaction with exon 8.
abstract: 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 26nt RNA from 5' end of exon 8, a 33nt RNA from 3' end of exon 10 and a 2225nt 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.
authors: School of life Sciences, Gwangju Institute of Science and Technology, Gwangju 500-712, Korea.
authors: Ha Na
authors: HN
authors: Jang
authors: School of life Sciences, Gwangju Institute of Science and Technology, Gwangju 500-712, Korea.
authors: Minho
authors: M
authors: Lee
authors: School of life Sciences, Gwangju Institute of Science and Technology, Gwangju 500-712, Korea.
authors: Tiing Jen
authors: TJ
authors: Loh
authors: School of life Sciences, Gwangju Institute of Science and Technology, Gwangju 500-712, Korea.
authors: Seung-Woo
authors: SW
authors: Choi
authors: School of life Sciences, Gwangju Institute of Science and Technology, Gwangju 500-712, Korea.
authors: Hyun Kyung
authors: HK
authors: Oh
authors: School of life Sciences, Gwangju Institute of Science and Technology, Gwangju 500-712, Korea.
authors: Heegyum
authors: H
authors: Moon
authors: School of life Sciences, Gwangju Institute of Science and Technology, Gwangju 500-712, Korea.
authors: Sunghee
authors: S
authors: Cho
authors: School of life Sciences, Gwangju Institute of Science and Technology, Gwangju 500-712, Korea.
authors: Seong-Eui
authors: SE
authors: Hong
authors: School of life Sciences, Gwangju Institute of Science and Technology, Gwangju 500-712, Korea.
authors: Do Han
authors: DH
authors: Kim
authors: Virginia Tech Carilion Research Institute, Roanoke, VA, 24016, USA; Department of Biomedical Sciences and Pathobiology, Virginia-Maryland Regional College of Veterinary Medicine, Virginia Tech, Blacksburg, VA, 24061, USA.
authors: Zhi
authors: Z
authors: Sheng
authors: Howard Hughes Medical Institute and Programs in Gene Function and Expression and Molecular Medicine, University of Massachusetts Medical School, Worcester, Massachusetts 01605, USA.
authors: Michael R
authors: MR
authors: Green
authors: School of life Sciences, Gwangju Institute of Science and Technology, Gwangju 500-712, Korea.
authors: Daeho
authors: D
authors: Park
authors: School of life Sciences, Gwangju Institute of Science and Technology, Gwangju 500-712, Korea.
authors: Xuexiu
authors: X
authors: Zheng
authors: School of life Sciences, Gwangju Institute of Science and Technology, Gwangju 500-712, Korea.
authors: Haihong
authors: H
authors: Shen
mesh_terms: D017398:Alternative Splicing / Q000235:genetics*; D017209:Apoptosis / Q000235:genetics*; D001665:Binding Sites / Q000235:genetics; D053143:Caspase 2 / Q000096:biosynthesis / Q000235:genetics*; D005091:Exons / Q000235:genetics; D055785:Gene Knockdown Techniques; D057809:HEK293 Cells; D006801:Humans; D016296:Mutagenesis; D012322:RNA Precursors / Q000235:genetics; D016601:RNA-Binding Proteins / Q000235:genetics* / Q000378:metabolism; D017384:Sequence Deletion; D000068103:Serine-Arginine Splicing Factors
chemical_list: D012322:RNA Precursors; D016601:RNA-Binding Proteins; C491388:SRSF3 protein, human; D000068103:Serine-Arginine Splicing Factors; D053143:Caspase 2
keywords: Anti-apoptotic; Apoptotic; Caspase-2; Exon 9; Pre-mRNA splicing; SRSF3