Manipulation of alternative splicing by a newly developed inhibitor of Clks
Manipulation of alternative splicing by a newly developed inhibitor of Clks
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DOI：
10.1074/jbc.m314298200
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发表时间：
2004-06-04
影响因子：
4.8
通讯作者：
Hagiwara, M
中科院分区：
文献类型：
--
作者：
Muraki, M;Ohkawara, B;Hagiwara, M
剪接位点使用的调节为控制基因表达和蛋白质组多样性的产生提供了一种通用机制，在许多生物过程中发挥着重要作用。越来越多的人类疾病归因于错误剪接事件，进一步说明了选择性剪接的重要性。剪接变体的适当空间和时间生成要求选择性剪接受到广泛的调控，类似于转录控制。 Clk（Cdc2 样激酶）家族与剪接控制有关，并且由至少四个成员组成。通过对化学库的广泛筛选，我们发现苯并噻唑化合物TG003对Clk1/Sty的活性具有有效的抑制作用。 TG003 通过抑制 Clk 介导的磷酸化，在体外抑制 β-珠蛋白前体 mRNA 的 SF2/ASF 依赖性剪接。该药物还抑制哺乳动物细胞中富含丝氨酸/精氨酸的蛋白质磷酸化、核斑点的解离以及 Clk1/Sty 依赖性选择性剪接。一致的是，给予 TG003 可以挽救爪蟾中由过度 Clk 活性引起的胚胎缺陷。因此，TG003这种Clk家族的新型抑制剂将成为剖析涉及体内富含丝氨酸/精氨酸的蛋白质磷酸化信号通路的调节机制的有价值的工具，并且可能适用于异常剪接的治疗操作。
The regulation of splice site usage provides a versatile mechanism for controlling gene expression and for the generation of proteome diversity, playing an essential role in many biological processes. The importance of alternative splicing is further illustrated by the increasing number of human diseases that have been attributed to mis-splicing events. Appropriate spatial and temporal generation of splicing variants demands that alternative splicing be subjected to extensive regulation, similar to transcriptional control. The Clk (Cdc2-like kinase) family has been implicated in splicing control and consists of at least four members. Through extensive screening of a chemical library, we found that a benzothiazole compound, TG003, had a potent inhibitory effect on the activity of Clk1/Sty. TG003 inhibited SF2/ASF-dependent splicing of beta-globin pre-mRNA in vitro by suppression of Clk-mediated phosphorylation. This drug also suppressed serine/arginine-rich protein phosphorylation, dissociation of nuclear speckles, and Clk1/Sty-dependent alternative splicing in mammalian cells. Consistently, administration of TG003 rescued the embryonic defects induced by excessive Clk activity in Xenopus. Thus, TG003, a novel inhibitor of Clk family will be a valuable tool to dissect the regulatory mechanisms involving serine/arginine-rich protein phosphorylation signaling pathways in vivo, and may be applicable for the therapeutic manipulation of abnormal splicing.