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

描述（由申请人提供）：肌动蛋白动力学的适当调节对于许多生物过程（如伤口愈合、免疫应答和形态发生）是必不可少的，并且在病理过程（如癌症转移和心血管疾病，发达国家的两大死亡原因）中起重要作用。尽管它广泛参与正常的生理和疾病，为治疗目的的目标肌动蛋白动力学的努力是在早期阶段，显然需要更深入地了解所涉及的过程。Arp 2/3复合物是肌动蛋白动力学中的关键参与者，其产生分支肌动蛋白阵列，其被认为对于许多细胞过程（包括细胞迁移、吞噬作用和细胞粘附）是重要的。使用来自条件性Arp 2/3基因敲除小鼠（Arpc 2基因）的细胞，我们提出解决肌动蛋白动力学领域的几个重要问题：1）Arp 2/3分支的肌动蛋白网络如何在细胞中分解和动态翻转？我们已经开发了一种新的光遗传学方法来控制Arp 2/3在细胞中的功能，这将使我们能够剖析去分支途径。2)Arp 2/3是肌动蛋白依赖的过程，如定向迁移，吞噬作用和细胞连接建立所必需的吗？这将使用干净的，在原代细胞中的基因缺失的方法，无论是离体使用活细胞成像方法和在体内使用多光子活体成像。3)细胞如何协调Arp 2/3和非Arp 2/3肌动蛋白途径以产生最佳的肌动蛋白动力学？使用我们的Arp 2/3缺陷的细胞，我们将询问Arp 2/3独立的途径，部分补偿其损失和研究Arp 2/3依赖和非依赖途径如何在协调的途径中发挥作用，以产生最佳的肌动蛋白在细胞中的动态。

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.