Beyond cell shape: Actin exerts systems-level control during morphogenesis

 描述(申请人提供)：细胞形状的改变，驱动形态发生和肌动蛋白缺乏会导致失败的细胞形状改变和出生缺陷。然而，我们对肌动蛋白对形态发生的影响的了解在很大程度上局限于分子清单和对大体表型的描述。虽然肌动蛋白细丝(F-肌动蛋白)被认为是细胞形状的主要结构决定因素，但基本问题仍然没有答案：F-肌动蛋白是如何重塑的？它是如何与细胞膜接触的？它是如何适应环境或遗传变异的？我们需要这些答案来理解发展和人类疾病。我们的长期目标是充分了解肌动蛋白在形成健康组织中的作用。我们关注的是果蝇的细胞化，这是在胚胎中建立第一层上皮膜的过程。在细胞化过程中，胚胎的每个新细胞都会发生三种细胞表面重塑事件：(I)微绒毛解体；(Ii)膜沟进入；(Iii)肌动球蛋白环收缩。我们已经证明，虽然这三个重塑事件在几十微米范围内是空间分离的，但它们仍然相互动力学地耦合在一起，并与肌动球蛋白环中可测量的变化相关联。我们还在肌动球蛋白环中确定了特定的F-肌动蛋白调节剂，它们促进细胞化对环境和遗传扰动的稳健性。这些结果表明，肌动蛋白不仅仅是赋予细胞形状。我们的工作模型是，基于肌动蛋白的机制也有助于形态发生的协调、动力学和稳健性。在这项拟议的工作中，我们将检验我们的模型如下：在目标1中，我们将检验一种假设，即由沟槽内倾产生的质膜张力抑制微绒毛中的F-肌动蛋白聚合，导致微绒毛“展开”或分离。在目标2中，我们将检验这一假说，即肌动球蛋白环收缩的不同动力学阶段受不同的F-肌动蛋白调节机制控制。在目标3中，我们将检验这样的假设：F-肌动蛋白调节剂sRYA和SPT稳定收缩环内的F-肌动蛋白以确保在 面对环境和遗传的扰动。我们已经组建了一个由物理学家、数学家和技术专家组成的多学科团队；我们将采用强大的方法组合，包括定量实时成像、体力分析、遗传学和基因组编辑，以不同寻常的细节研究细胞化事件。由于驱动细胞化的基于肌动蛋白的机制被广泛保守，我们的发现将与包括人类在内的其他器官中毒相关。我们将确定单个重塑事件是如何在空间和时间上进行调节的，同时还将展示这些事件如何相互协调，以实现高保真的细胞化。在这项工作中，我们还将揭示基因、机制和环境如何调节F-肌动蛋白。

 DESCRIPTION (provided by applicant): Changes in cell shape drive morphogenesis and actin deficiencies cause failed cell shape change and birth defects. Yet our knowledge of actin's influence on morphogenesis is largely limited to lists of molecules and descriptions of gross phenotypes. While actin filaments (F-actin) are recognized as the major structural determinant of cell shape, basic questions remain unanswered: How is F-actin remodeled? How does it inter- face with cell membranes? How does it adapt to environmental or genetic variability? We need these answers to understand development and human disease. Our long-term goal is to fully understand actin's role in forming healthy tissues. We are focused on Drosophila cellularization, the process that builds the first epithelial sheet in the embryo. During cellularization, three cell surface remodeling events are going on in every new cell of the embryo: (i) microvilli disassemble; (ii) a membrane furrow ingresses; and (iii) an actomyosin ring contracts. We have shown that while these three remodeling events are spatially segregated over tens of microns, they are still kinetically coupled to each other and to measurable changes in the actomyosin ring. We have also identified specific F-actin regulators in the actomyosin ring that promote the robustness of cellularization against environmental and genetic perturbation. These results argue that actin does more than just give cells their shape. Our working model is that actin- based mechanisms also contribute to the coordination, kinetics, and robustness of morphogenesis. In this proposed work, we will test our model as follows: In Aim 1, we will test the hypothesis that plasma membrane tension, generated by furrow ingression, suppresses F-actin polymerization in microvilli, causing them to "unfold" or fall apart. In Aim 2, we will test he hypothesis that distinct kinetic phases of actomyosin ring contraction are governed by distinct mechanisms of F-actin regulation. In Aim 3, we will test the hypothesis that F-actin regulators, Srya and Spt, stabilize F-actin within the contractile ring to ensure robust cellularization in the face of environmental and genetic perturbation. We have assembled a multidisciplinary team of physicists, mathematicians and technologists; and we will employ a powerful combination of approaches, including quantitative live imaging, physical force assays, genetics, and genome editing to examine the events of cellularization in uncommon detail. Because the actin- based machinery that drives cellularization is widely conserved, our findings will be relevant to other organ- isms, including humans. We will establish how individual remodeling events are regulated in space and time, while also showing how the events are coordinated with each other to accomplish cellularization at high fidelity. In doing this work, we will also reveal how genes, mechanics, and environment regulate F-actin.