β-Arrestin 1 Inhibits the GTPase-Activating Protein Function of ARHGAP21, Promoting Activation of RhoA following Angiotensin II Type 1A Receptor Stimulation
β-Arrestin 1 Inhibits the GTPase-Activating Protein Function of ARHGAP21, Promoting Activation of RhoA following Angiotensin II Type 1A Receptor Stimulation
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DOI：
10.1128/mcb.00883-10
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发表时间：
2011-03-01
影响因子：
5.3
通讯作者：
Baillie, G. S.
中科院分区：
文献类型：
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作者：
Anthony, D. F.;Sin, Y. Y.;Baillie, G. S.
已知血管紧张素 II 刺激后小 GTP 酶 RhoA 的激活会导致肌动蛋白重组和应力纤维形成。血管紧张素 II 完全激活 RhoA 取决于支架蛋白 β-arrestin 1，尽管其背后的机制仍不清楚。在这里，我们发现了 β-arrestin 1 的新伙伴和功能，即与 ARHGAP21（也称为 ARHGAP10）结合，ARHGAP21 是一种已知的 RhoA 活性效应子，它抑制其 GTP 酶激活蛋白（GAP）功能。使用酵母双杂交筛选、肽阵列、体外结合研究、截短分析和免疫共沉淀技术，我们证明 β-arrestin 1 在横断 RhoA 效应器 GAP 结构域的区域中直接与 ARHGAP21 结合。此外，我们发现，含有 β-arrestin 1 和 ARHGAP21 的复合物的水平在血管紧张素刺激后动态增加，并且这种相互作用的动力学调节 RhoA 的时间激活。利用从肽阵列中收集的信息，我们开发了一种细胞渗透肽，可以抑制这些蛋白质的相互作用。使用这种肽，我们证明β-抑制蛋白1/ARHGAP21复合物的破坏会产生更活跃的ARHGAP21，从而导致通过血管紧张素II 1A型受体的信号传导效率较低，从而减弱受刺激的应力纤维形成。
Activation of the small GTPase RhoA following angiotensin II stimulation is known to result in actin reorganization and stress fiber formation. Full activation of RhoA, by angiotensin II, depends on the scaffolding protein beta-arrestin 1, although the mechanism behind its involvement remains elusive. Here we uncover a novel partner and function for beta-arrestin 1, namely, in binding to ARHGAP21 (also known as ARHGAP10), a known effector of RhoA activity, whose GTPase-activating protein (GAP) function it inhibits. Using yeast two-hybrid screening, a peptide array, in vitro binding studies, truncation analyses, and coimmunoprecipitation techniques, we show that beta-arrestin 1 binds directly to ARHGAP21 in a region that transects the RhoA effector GAP domain. Moreover, we show that the level of a complex containing beta-arrestin 1 and ARHGAP21 is dynamically increased following angiotensin stimulation and that the kinetics of this interaction modulates the temporal activation of RhoA. Using information gleaned from a peptide array, we developed a cell-permeant peptide that serves to inhibit the interaction of these proteins. Using this peptide, we demonstrate that disruption of the beta-arrestin 1/ARHGAP21 complex results in a more active ARHGAP21, leading to less-efficient signaling via the angiotensin II type 1A receptor and, thereby, attenuation of stimulated stress fiber formation.