Regulation of actin during cell migration

项目摘要 细胞迁移驱动组织形态发生、免疫反应和癌症等关键生物学事件 转移瘤。我们最近的数据令人惊讶地表明，细胞前沿的形成和功能在 迁移关键依赖于精氨酸化，这是一种相对未被探索的翻译后修饰。此外，在 基于细胞的表型挽救实验，我们已经证明了实验上精氨酸化的β肌动蛋白可以 小鼠胚胎成纤维细胞缺乏精氨酸酶后细胞前沿功能的恢复 AT1。在我们正在进行的研究中，确定β-肌动蛋白的N-末端精氨酸化如何与片状脂血症有关 形成和定向细胞迁移，我们已经做出了新的观察结果，即 精氨酸化的β肌动蛋白在迁移过程中以细胞前沿为靶点。此外，我们的数据表明 β-肌动蛋白的精氨酸化受其mRNA序列的选择性调控，而不是其蛋白质结构。在……里面 在氨基酸水平上与β-肌动蛋白有99%同源性的支撑体-肌动蛋白-肌动蛋白 序列。我们已经证明，这种差异直接导致β肌动蛋白和β肌动蛋白的翻译速度更快 通过一种与蛋白质泛素化耦合的新机制，导致其选择性精氨酸化。它也是众所周知的 Zipcode介导的β肌动蛋白mRNA靶向调节其前沿定位，与精氨酸化一样， 对细胞定向迁移是必不可少的。我们假设信使核糖核酸介导的N-末端调节 β-肌动蛋白的精氨酸化通过促进肌动蛋白独特地调节细胞迁移过程中的肌动蛋白功能 在细胞前沿聚合。在这个提案中，我们将通过三个具体目标来检验这一假设 这将：(1)检验β-肌动蛋白精氨酸化促进细胞内肌动蛋白聚合的假设。 (2)检验β-肌动蛋白功能受编码和非编码区唯一调控的假设 它的mrna；以及(3)检验这种调节与细胞内精氨酸化的调节相耦合的假设。 细胞迁移过程中的活动。这些实验将解决控制细胞的一种新的调控机制 通过调节肌动蛋白的属性和mRNA结构的极化和运动性，最终将使 对细胞在基本生理事件中迁移的定向功能研究达到了新的水平。

Project Summary Cell migration drives such key biological events as tissue morphogenesis, immune response, and cancer metastases. Our recent data surprisingly show that the formation and function of the cell leading edge during migration critically depends on arginylation, a relatively unexplored posttranslational modification. Moreover, in cell-based phenotype rescue experiments, we have shown that experimentally arginylated beta actin can largely restore cell leading edge function in mouse embryonic fibroblasts lacking the arginylation enzyme ATE1. In our ongoing studies to determine how N-terminal arginylation of beta actin contributes to lamellipodia formation and directed cell migration, we have made the novel observations that a prominent subset of arginylated beta actin is targeted to the cell leading edge during migration. Moreover, our data demonstrate that beta actin arginylation is selectively regulated by its mRNA sequence rather than its protein structure. In support, gamma actin, which is 99% identical to beta actin at the amino acid level, differs by 13% in its mRNA sequence. We have shown that this difference is directly responsible for faster translation rate of beta actin and leads to its selective arginylation through a novel mechanism coupled to protein ubiquitination. It is also known that zipcode-mediated beta actin mRNA targeting regulates its leading edge localization and, like arginylation, is essential for directional cell migration. We hypothesize that mRNA-mediated regulation of N-terminal arginylation of beta actin uniquely regulates actin function during cell migration by facilitating actin polymerization at the cell leading edge. In this proposal, we will test this hypothesis through three specific aims that will: (1) test the hypothesis that beta actin arginylation facilitates actin polymerization at the cell leading egde; (2) test the hypothesis that beta actin function is uniquely regulated by coding and noncoding regions of its mRNA; and (3) test the hypothesis that this regulation is coupled to modulation of intracellular arginylation activity during cell migration. These experiments will address a novel regulatory mechanism controlling cell polarization and motility through modulating actin's properties and mRNA structure and will ultimately enable a new level of targeted functional studies of cell migration during essential physiological events.