Morphogenesis: Biophysics and Genetics of Dorsal Closure

描述（由申请方提供）：果蝇的背侧闭合是发育和伤口愈合期间细胞片形态发生的模型系统。我们计划调查分子，细胞，和新兴的属性，推动形态在关闭过程中使用生物物理（激光显微外科），遗传，药理学和建模方法。以前，我们表明，非肌肉肌球蛋白II驱动的浆膜和超细胞，肌动球蛋白丰富的荷包，这两个组织的收缩性驱动的大部分进展关闭。此外，我们表明，组织力是由粘附介导的拉链协调的，前后拉链速率常数之间存在不对称性，并且拉链速率常数可以响应于激光扰动而上调。我们还表明，去除一种或另一种力会导致剩余力的上调，并且以野生型速率完成闭合。此外，浆膜张力和拉链速率常数的上调一起解决了闭合的稳健性和弹性。最后，我们证明了驱动闭合的力的矢量和比贡献的单个力小两到三个数量级。这表明，需要调节这些大的力量，使细胞片无情地关闭。在这里，我们专注于应用激光手术，药理学和定量建模工具，探索新兴的属性，是驱动野生型和突变动物细胞片形态发生的细胞和分子机器的后果。通过将这些方法应用于野生型胚胎和在闭合的一个或另一个方面失败的选定突变胚胎的分析，我们计划以下内容。1)我们将调查的假设，机械门控通道和/或细胞基质和细胞细胞连接的感觉和响应的力量，以调节关闭的速度。2)我们将测量每种组织闭合所需力的绝对大小。3）我们将研究微管在调节肌动蛋白关闭功能中的作用。4)我们将制定数学模型，概括在组织和/或细胞分辨率的行为关闭突变，微扰或激光研究胚胎。果蝇细胞片形态发生的这些研究将为协调脊椎动物形态发生和伤口愈合中细胞形状变化的生物学过程提供细胞和分子基础。公共卫生相关性这项工作的重点是背部闭合，果蝇黑腹果蝇的一个过程，模拟脊椎动物的细胞片运动。果蝇为多学科方法提供了独特的机会，许多参与运动的蛋白质在果蝇和人类之间高度保守（有些蛋白质具有> 90%的相同性，许多人类蛋白质可以通过实验挽救其果蝇直系同源物中的遗传缺陷）。此外，可比较的床单运动表征了人类发育（例如，神经管形成）和伤口愈合的早期阶段。此外，在闭合期间发生的细胞-细胞连接和细胞-基质连接之间的程序化相互作用对于表皮-间充质转化至关重要，当上皮细胞转移并导致癌症时，表皮-间充质转化被激活。在果蝇中的这些研究将提供对运动的分子和细胞基础的深入了解，并提供一个窗口，以表征形态发生和伤口愈合的紧急行为。

DESCRIPTION (provided by applicant): Drosophila's Dorsal Closure is a model system for cell sheet morphogenesis during development and wound healing. We plan to investigate the molecular, cellular, and emergent properties that drive morphogenesis during closure using biophysical (laser microsurgery), genetic, pharmacological and modeling approaches. Previously, we showed that nonmuscle myosin II drives contractility in the amnioserosa and in the supracellular, actomyosin-rich purse-strings and that both tissues drive the bulk of progress toward closure. In addition we showed that the tissue forces are coordinated by adhesion- mediated zipping, that there is an asymmetry between the anterior and posterior zipping rate constants, and that the zipping rate constant can be upregulated in response to laser perturbation. We also showed that the removal of one or another force leads to upregulation of the forces that remain and closure proceeds to completion at wild type rates. Furthermore, the upregulation of the amnioserosa force and the zipping rate constant together address the robustness and resiliency of closure. Finally, we showed that the vector sum of the forces that drive closure is two to three orders of magnitude smaller than the individual forces that contribute. This indicates that regulation of these large forces is required so that cell sheets move inexorably to closure. Here we focus on applying laser-surgical, pharmacological and quantitative-modeling tools to explore the emergent properties that are the consequence of the cellular and molecular machines that drive cell sheet morphogenesis in wild type and mutant animals. By applying these methods to the analysis of wild type embryos an selected mutant embryos that fail in on or another aspect of closure, we plan the following. 1) We will investigate the hypotheses that mechanically gated channels and/or cell-matrix and cell-cell junctions sense and respond to forces to regulate the rate of closure. 2) We will measure the absolute magnitude of the forces each tissue contributes to closure. 3)We will investigate the role of microtubules in regulating actin function for closure. 4) We will formulate mathematical models that recapitulate at tissue and/or cellular resolution the behavior of closure in mutant, pharmacologically perturbed or laser investigated embryos. These studies on cell sheet morphogenesis in Drosophila will provide insight into the cellular and molecular basis for the biological processes that coordinate cell shape changes in vertebrate morphogenesis and wound healing. PUBLIC HEALTH RELEVANCE This work focuses on dorsal closure, a process in the fruit fly Drosophila melanogaster that models cell sheet movements in vertebrates. Drosophila offers unique opportunities for multidisciplinary approaches and many of the proteins involved in movement are highly conserved between flies and humans (some are > 90% identical and many human proteins can experimentally rescue genetic defects in their fly orthologs). In addition, comparable sheet movements characterize early stages of human development (for example, neural tube formation) and wound healing. Moreover, the programmed interplay between cell-cell junctions and cell-matrix junctions that occurs during closure is crucial for the epidermal-mesenchymal transition that is activated when epithelial cells become metastatic and cause cancer. These studies in Drosophila will provide insight into the molecular and cellular basis of motility and provide a window onto the emergent behaviors that characterize morphogenesis and wound healing.