Humphries Christine L, Balcer Heath I, D'Agostino Jessica L, Winsor Barbara, Drubin David G, Barnes Georjana, Andrews Brenda J, Goode Bruce L
Department of Molecular and Medical Genetics, University of Toronto, Ontario M5S 1A8, Canada.
J Cell Biol. 2002 Dec 23;159(6):993-1004. doi: 10.1083/jcb.200206113.
Mechanisms for activating the actin-related protein 2/3 (Arp2/3) complex have been the focus of many recent studies. Here, we identify a novel mode of Arp2/3 complex regulation mediated by the highly conserved actin binding protein coronin. Yeast coronin (Crn1) physically associates with the Arp2/3 complex and inhibits WA- and Abp1-activated actin nucleation in vitro. The inhibition occurs specifically in the absence of preformed actin filaments, suggesting that Crn1 may restrict Arp2/3 complex activity to the sides of filaments. The inhibitory activity of Crn1 resides in its coiled coil domain. Localization of Crn1 to actin patches in vivo and association of Crn1 with the Arp2/3 complex also require its coiled coil domain. Genetic studies provide in vivo evidence for these interactions and activities. Overexpression of CRN1 causes growth arrest and redistribution of Arp2 and Crn1p into aberrant actin loops. These defects are suppressed by deletion of the Crn1 coiled coil domain and by arc35-26, an allele of the p35 subunit of the Arp2/3 complex. Further in vivo evidence that coronin regulates the Arp2/3 complex comes from the observation that crn1 and arp2 mutants display an allele-specific synthetic interaction. This work identifies a new form of regulation of the Arp2/3 complex and an important cellular function for coronin.
激活肌动蛋白相关蛋白2/3（Arp2/3）复合物的机制一直是近期许多研究的焦点。在此，我们鉴定出一种由高度保守的肌动蛋白结合蛋白冠蛋白介导的Arp2/3复合物调节新模式。酵母冠蛋白（Crn1）与Arp2/3复合物发生物理结合，并在体外抑制WA和Abp1激活的肌动蛋白成核作用。这种抑制作用在不存在预先形成的肌动蛋白丝时特异性发生，这表明Crn1可能将Arp2/3复合物的活性限制在丝的侧面。Crn1的抑制活性存在于其卷曲螺旋结构域中。Crn1在体内定位于肌动蛋白斑点以及Crn1与Arp2/3复合物的结合也需要其卷曲螺旋结构域。遗传学研究为这些相互作用和活性提供了体内证据。CRN1的过表达导致生长停滞以及Arp2和Crn1p重新分布到异常的肌动蛋白环中。这些缺陷可通过缺失Crn1卷曲螺旋结构域以及通过Arp2/3复合物p35亚基的等位基因arc35 - 26来抑制。进一步的体内证据表明冠蛋白调节Arp2/3复合物，这来自于观察到crn1和arp2突变体表现出等位基因特异性的合成相互作用。这项工作鉴定出了Arp2/3复合物调节的一种新形式以及冠蛋白的一项重要细胞功能。
