The role of the Arp2/3 complex in cellular actin dynamics

描述(申请人提供)：肌动蛋白动力学的适当调节对伤口愈合、免疫反应和形态形成等许多生物学过程至关重要，并在癌症转移和心血管疾病等病理过程中发挥重要作用，癌症转移和心血管疾病是发达国家的两大死亡原因。尽管肌动蛋白广泛参与正常生理和疾病，但为了治疗目的而针对肌动蛋白动力学的努力还处于早期阶段，显然需要对涉及的过程有更深入的了解。Arp2/3复合体是肌动蛋白动力学中的关键角色，它产生分支肌动蛋白阵列，被认为对许多细胞过程至关重要，包括细胞迁移、吞噬和细胞黏附。使用来自条件Arp2/3基因敲除小鼠(Arpc2基因)的细胞，我们建议解决肌动蛋白动力学领域的几个重要问题：1)Arp2/3分支肌动蛋白网络是如何在细胞中分解和动态翻转的？我们已经开发了一种新的光遗传学方法来控制细胞中的Arp2/3功能，这将使我们能够剖析去分支途径。2)肌动蛋白依赖的过程，如定向迁移、吞噬和细胞-细胞连接的建立是否需要Arp2/3？这将通过在体外使用活细胞成像方法在原代细胞中使用干净的基因缺失方法来解决，并使用体内使用多光子活体成像来解决。3)细胞如何协调Arp2/3和非Arp2/3肌动蛋白通路以产生最佳的肌动蛋白动力学？利用我们的Arp2/3缺陷细胞，我们将询问部分补偿Arp2/3缺失的Arp2/3非依赖途径，并研究Arp2/3依赖和非依赖途径如何在细胞中产生最佳的肌动蛋白动力学。

DESCRIPTION (provided by applicant): The proper regulation of actin dynamics is essential for many biological processes such as wound healing, immune response and morphogenesis, and plays a significant role in pathological processes such as cancer metastasis and cardiovascular disease, the two leading causes of death in the developed world. Despite its widespread involvement in normal physiology and disease, efforts to target actin dynamics for therapeutic purposes are at an early stage and clearly require a deeper understanding of the processes involved. The Arp2/3 complex is a critical player in actin dynamics that generates branched actin arrays which are thought to be important for many cellular processes including cell migration, phagocytosis and cell adhesion. Using cells derived from a conditional Arp2/3 knockout mouse (Arpc2 gene), we propose to address several important questions for the field of actin dynamics: 1) How are Arp2/3-branched actin networks disassembled and dynamically turned over in cells? We have developed a new optogenetic method to control Arp2/3 function in cells with light that will allow us to dissect the de-branching pathway. 2) Is Arp2/3 required for actin-dependent processes such as directed migration, phagocytosis and cell-cell junction establishment? This will be addressed using a clean, genetic deletion approach in primary cells both ex vivo using live-cell imaging approaches and in vivo using multiphoton intravital imaging. 3) How do cells coordinate Arp2/3 and non-Arp2/3 actin pathways to produce optimal actin dynamics? Using our Arp2/3- deficient cells, we will interrogate the Arp2/3-independent pathways that partially compensate for its loss and study how Arp2/3-dependent and -independent pathway act in a coordinated pathway to produce optimal actin dynamics in cells.