Three regions within ActA promote Arp2/3 complex-mediated actin nucleation and Listeria monocytogenes motility.
Three regions within ActA promote Arp2/3 complex-mediated actin nucleation and Listeria monocytogenes motility.
复制标题
ACTA中的三个区域促进了ARP2/3复合介导的肌动蛋白成核和单核细胞增生李斯特菌的运动。
DOI：
10.1083/jcb.150.3.527
复制
发表时间：
2000-08-07
影响因子：
7.8
通讯作者：
Welch, M D
中科院分区：
文献类型：
--
作者：
Skoble, J;Portnoy, D A;Welch, M D
单核增生李斯特菌ActA蛋白通过增强宿主Arp2/3复合体的肌动蛋白成核活性来诱导肌动蛋白为基础的运动。通过系统截断分析，我们确定了一个136个nh2末端片段，该片段在体外刺激成核中完全活跃。进一步的缺失分析表明，该片段包含三个区域，它们对成核很重要，与宿主WASP家族蛋白具有功能和/或有限的序列相似性：酸性拉伸区、肌动蛋白单体结合区和cofilin同源序列。为了确定每个区域对肌动蛋白运动的贡献，我们将ActA衍生物的生化活性与细胞中相应突变菌的表型进行了比较。酸性拉伸的作用是提高肌动蛋白成核的效率，运动的速度和频率，以及细胞-细胞扩散的有效性。在体外，肌动蛋白成核需要单体结合区，但在感染细胞中肌动蛋白聚合或运动不需要单体结合区，这表明在宿主细胞质中招募单体可能存在冗余机制。cofilin同源序列是体外刺激肌动蛋白与Arp2/3复合物成核的关键，也是细胞内肌动蛋白聚合和运动的必要条件。这些数据表明，每个区域都参与基于肌动蛋白的运动，并且cofilin同源序列在Arp2/3复合物的激活中起主要作用，并且是单核增生乳杆菌发病机制的重要决定因素。
The Listeria monocytogenes ActA protein induces actin-based motility by enhancing the actin nucleating activity of the host Arp2/3 complex. Using systematic truncation analysis, we identified a 136-residue NH2-terminal fragment that was fully active in stimulating nucleation in vitro. Further deletion analysis demonstrated that this fragment contains three regions, which are important for nucleation and share functional and/or limited sequence similarity with host WASP family proteins: an acidic stretch, an actin monomer–binding region, and a cofilin homology sequence. To determine the contribution of each region to actin-based motility, we compared the biochemical activities of ActA derivatives with the phenotypes of corresponding mutant bacteria in cells. The acidic stretch functions to increase the efficiency of actin nucleation, the rate and frequency of motility, and the effectiveness of cell–cell spread. The monomer-binding region is required for actin nucleation in vitro, but not for actin polymerization or motility in infected cells, suggesting that redundant mechanisms may exist to recruit monomer in host cytosol. The cofilin homology sequence is critical for stimulating actin nucleation with the Arp2/3 complex in vitro, and is essential for actin polymerization and motility in cells. These data demonstrate that each region contributes to actin-based motility, and that the cofilin homology sequence plays a principal role in activation of the Arp2/3 complex, and is an essential determinant of L. monocytogenes pathogenesis.