Relative strength of 5’ splice-site strength defines functions of SRSF2 and SRSF6 in alternative splicing of Bcl-x pre-mRNA |
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Authors: | Namjeong Choi Yongchao Liu Jagyeong Oh Jiyeon Ha Claudia Ghigna Xuexiu Zheng Haihong Shen |
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Institution: | 1.School of Life Sciences, Gwangju Institute of Science and Technology, Gwangju 61005, Korea;2.Institute of Molecular Genetics “Luigi Luca Cavalli-Sforza”, National Research Council, Pavia 27100, Italy |
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Abstract: | Bcl-x, a member of the Bcl-2 family, plays a key role in apoptosis. Alternative splicing of Bcl-x pre-mRNA through alternative 5’ splice-site selection produces an anti-apoptotic mRNA isoform that includes exon 2b and a pro-apoptotic Bcl-x mRNA isoform that excludes exon 2b. Here we used Bcl-x minigene and identified SRSF2 and SRSF6 as two regulatory factors of 5’ splice-site selection of Bcl-x pre-mRNA. We selected binding clusters closer to 5’ splice-sites from multiple potential binding sites of SRSF2 and SRSF6 to perform loss of functions analysis through site-directed mutagenesis. Our results demonstrated that these mutations did not abolish regulatory functions of SRSF2 or SRSF6, indicating that a single binding motif or a cluster was not a functional target of these proteins in Bcl-x pre-mRNA splicing. Random deletion mutagenesis did not disrupt the role of SRSF2 and SRSF6. Importantly, mutagenesis of 5’ splice-site to a conserved or a weaker score demonstrated that the weaker strength of the target 5’ splice-site or higher strength of the other 5’ splice-site strength limited the role of SRSF2 and SRSF6 in 5’ splice-site activation. |
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Keywords: | 5’ splice-site Alternative splicing Bcl-x SRSF2 SRSF6 |
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