MBNL and CELF proteins regulate alternative splicing of the skeletal muscle chloride channel CLCN1
MBNL and CELF proteins regulate alternative splicing of the skeletal muscle chloride channel CLCN1
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DOI：
10.1093/nar/gkp681
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发表时间：
2009-10-01
影响因子：
14.9
通讯作者：
Ishiura, Shoichi
中科院分区：
文献类型：
--
作者：
Kino, Yoshihiro;Washizu, Chika;Ishiura, Shoichi
骨骼肌氯离子通道CLCN 1/ClC-1的表达和功能受选择性剪接调控。CLCN 1外显子7A的包含在强直性肌营养不良（DM）中异常升高，DM是一种由CTG或CCTG重复扩增引起的遗传性疾病。外显子7A包含增加导致CLCN 1功能降低，这可能是肌强直的原因。两种RNA结合蛋白家族--肌盲样蛋白（MBNL）和CUG-BP和ETR-3样因子（CELF）蛋白--被认为介导DM中的剪接失调。在这里，我们已经确定了多种因素，调节小鼠Clcn 1小基因的选择性剪接。外显子7A的包含被MBNL蛋白抑制，而被扩展的CUG重复序列或CELF 4促进，但不被CUG-BP促进。突变分析表明，外显子7A及其侧翼区域介导MBNL 1的作用，而内含子6中的另一个不同区域介导CELF 4的作用。在外显子7A的5'端发现了一个外显子剪接增强子，该外显子剪接增强子对外显子7A的包含至关重要，可能被MBNL 1抑制。总的来说，这些结果提供了Clcn 1剪接的调节机制模型，并揭示了MBNL和CELF蛋白的新的调节特性。
The expression and function of the skeletal muscle chloride channel CLCN1/ClC-1 is regulated by alternative splicing. Inclusion of the CLCN1 exon 7A is aberrantly elevated in myotonic dystrophy (DM), a genetic disorder caused by the expansion of a CTG or CCTG repeat. Increased exon 7A inclusion leads to a reduction in CLCN1 function, which can be causative of myotonia. Two RNA-binding protein families-muscleblind-like (MBNL) and CUG-BP and ETR-3-like factor (CELF) proteins-are thought to mediate the splicing misregulation in DM. Here, we have identified multiple factors that regulate the alternative splicing of a mouse Clcn1 minigene. The inclusion of exon 7A was repressed by MBNL proteins while promoted by an expanded CUG repeat or CELF4, but not by CUG-BP. Mutation analyses suggested that exon 7A and its flanking region mediate the effect of MBNL1, whereas another distinct region in intron 6 mediates that of CELF4. An exonic splicing enhancer essential for the inclusion of exon 7A was identified at the 5' end of this exon, which might be inhibited by MBNL1. Collectively, these results provide a mechanistic model for the regulation of Clcn1 splicing, and reveal novel regulatory properties of MBNL and CELF proteins.