hnRNP H enhances skipping of a nonfunctional exon P3A in CHRNA1 and a mutation disrupting its binding causes congenital myasthenic syndrome
hnRNP H enhances skipping of a nonfunctional exon P3A in CHRNA1 and a mutation disrupting its binding causes congenital myasthenic syndrome
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DOI：
10.1093/hmg/ddn305
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发表时间：
2008-12-15
影响因子：
3.5
通讯作者：
Ohno, Kinji
中科院分区：
文献类型：
--
作者：
Masuda, Akio;Shen, Xin-Ming;Ohno, Kinji
在人类和类人猿中，编码肌肉烟碱乙酰胆碱受体α亚基的CHRNA 1携带一个框内外显子P3 A，包含该外显子产生一个无功能的α亚基。在肌肉中，P3 A（-）和P3 A（+）转录物以1：1的比例产生，但选择性剪接的功能意义和调控仍然难以捉摸。在患有先天性肌无力综合征的患者中鉴定的内含子突变（IVS 3 - 8 G> A）破坏内含子剪接沉默子（ISS）并导致下游P3 A外显子的排他性包含。我们发现ISS结合剪接反式因子是异质核核糖核蛋白（hnRNP）H，突变使hnRNP对ISS的亲和力减弱了近100倍。我们接下来表明，hnRNP H直接放置到内含子3沉默的3'端，并且siRNA介导的hnRNP H下调增强了外显子P3 A的识别。对人类基因组的分析表明，hnRNPH结合UGGG基序在内含子的3'端附近过度表达。根据这一线索，我们发现GRIP 1、FAS、VPS 13 C和NRCAM的替代外显子被hnRNP H下调。我们的研究结果表明，hnRNP H-结合基序接近内含子的3'端的存在是一个重要的，但低估了下游外显子的剪接调节。
In humans and great apes, CHRNA1 encoding the muscle nicotinic acetylcholine receptor alpha subunit carries an inframe exon P3A, the inclusion of which yields a nonfunctional alpha subunit. In muscle, the P3A(-) and P3A(+) transcripts are generated in a 1:1 ratio but the functional significance and regulation of the alternative splicing remain elusive. An intronic mutation (IVS3-8G > A), identified in a patient with congenital myasthenic syndrome, disrupts an intronic splicing silencer (ISS) and results in exclusive inclusion of the downstream P3A exon. We found that the ISS-binding splicing trans-factor was heterogeneous nuclear ribonucleoprotein (hnRNP) H and the mutation attenuated the affinity of hnRNP for the ISS similar to 100-fold. We next showed that direct placement of hnRNP H to the 3' end of intron 3 silences, and siRNA-mediated downregulation of hnRNP H enhances recognition of exon P3A. Analysis of the human genome suggested that the hnRNPH-binding UGGG motif is overrepresented close to the 3' ends of introns. Pursuing this clue, we showed that alternative exons of GRIP1, FAS, VPS13C and NRCAM are downregulated by hnRNP H. Our findings imply that the presence of the hnRNP H-binding motif close to the 3' end of an intron is an essential but underestimated splicing regulator of the downstream exon.