Regulation of Rnd3 localization and function by protein kinase C alpha-mediated phosphorylation.
Regulation of Rnd3 localization and function by protein kinase C alpha-mediated phosphorylation.
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DOI：
10.1042/bj20082377
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发表时间：
2009-10-23
期刊：
影响因子：
--
通讯作者：
Cox AD
中科院分区：
文献类型：
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作者：
Madigan JP;Bodemann BO;Brady DC;Dewar BJ;Keller PJ;Leitges M;Philips MR;Ridley AJ;Der CJ;Cox AD
Rnd蛋白（Rnd1， Rnd2和Rnd3/RhoE）形成了Rho小gtpase家族的一个独特分支。Rnd3表达的改变导致细胞骨架组织和细胞周期进程的改变。Rnd3的功能是降低RhoA活性，但Rnd3本身是如何被调节以引起这些变化的仍在研究中。与其他Rho家族蛋白不同，Rnd3不受GTP/GDP循环的调节，而是在表达水平和翻译后修饰（如戊烯酰化和磷酸化）的调节。我们在这里表明，在PKC激动剂刺激下，Rnd3经历了电泳迁移，其亚细胞定位在细胞膜上变得丰富。这些变化被常规PKC异构体的抑制所阻断，并且不会发生在PKCα-缺失细胞或Rnd3的非磷酸化突变体中。我们进一步证明PKCα在体外激酶试验中直接磷酸化Rnd3。此外，我们提供的证据表明，Rnd3的磷酸化状态对其阻断Rho-ROCK通路信号传导的能力有直接影响。这些结果确定了另一种调节机制，并澄清了Rnd3如何调节Rho信号以改变细胞骨架组织。
The Rnd proteins (Rnd1, Rnd2 and Rnd3/RhoE) form a distinct branch of the Rho family of small GTPases. Altered Rnd3 expression causes changes in cytoskeletal organization and cell cycle progression. Rnd3 functions to decrease RhoA activity, but how Rnd3 itself is regulated to cause these changes is still under investigation. Unlike other Rho family proteins, Rnd3 is regulated not by GTP/GDP cycling, but at the level of expression and by posttranslational modifications such as prenylation and phosphorylation. We show here that, upon PKC agonist stimulation, Rnd3 undergoes an electrophoretic mobility shift and its subcellular localization becomes enriched at internal membranes. These changes are blocked by inhibition of conventional PKC isoforms and do not occur in PKCα-null cells or to a nonphosphorylatable mutant of Rnd3. We further show that PKCα directly phosphorylates Rnd3 in an in vitro kinase assay. Additionally, we provide evidence that the phosphorylation status of Rnd3 has a direct effect on its ability to block signaling from the Rho-ROCK pathway. These results identify an additional mechanism of regulation and provide clarification of how Rnd3 modulates Rho signaling to alter cytoskeletal organization.