Cellular Remodeling by Microtubule Severing

通过微管切断进行细胞重塑

• 批准号: 8667814
• 负责人: JENNIFER L ROSS
• 金额: $ 30.22万
• 依托单位: ALBERT EINSTEIN COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-07-01 至 2018-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8667814
• 关键词: ATP phosphohydrolase; Actins; Animal Model; Basic Science; Biophysical Process; Biophysics; Cardiovascular Diseases; Cell Polarity; Cell division; Cell model; Cell physiology; Cells; Cellular Assay; Centrosome; Clinical; Collaborations; Complex; Cytoskeleton; Data; Development; Disease; Enzymes; Etiology; Excision; Generations; Goals; Health; Human; Human Development; Image Analysis; In Vitro; Injury; Knowledge; Life; Maintenance; Malignant Neoplasms; Mammalian Cell; Mammals; Massachusetts; Medicine; Membrane; Mental Retardation; Methods; Microtubule Proteins; Microtubules; Minus End of the Microtubule; Modeling; Molecular; Morphogenesis; Movement; Names; Normal Cell; Paper; Pathology; Physics; Physiology; Positioning Attribute; Process; Proteins; Published Comment; Publishing; Regulation; Research; Research Proposals; Role; Signal Transduction; Somatic Cell; Testing; Testis; Translating; Tubulin; Universities; Work; Wound Healing; base; cell motility; cell type; cellular imaging; college; in vivo; insight; katanin; member; neovascularization; novel; novel therapeutics; polymerization; prevent; professor; programs; public health relevance; reconstitution; repaired; response; single molecule; spastin; tissue regeneration; tissue repair

DESCRIPTION (provided by applicant): Rapid remodeling of the microtubule cytoskeleton is essential for normal cell division, motility and morphogenesis. A unique and intriguing class of proteins involved in this remodeling are the microtubule severing enzymes, so named because of their ability to generate internal breaks in the microtubule lattice, in vitro. The studies outlned in this research proposal will elucidate the cellular functions and biophysical mechanisms of action of members of a still poorly understood subfamily of microtubule severing enzymes, termed fidgetins. The founding member of this subfamily, fidgetin, has long been known to be important for mammalian development, yet the mechanistic basis of its developmental functions remains unclear. In a recent study, we showed that human fidgetin is a microtubule severing enzyme and minus-end depolymerase. We have now found that fidgetin and the closely related protein fidgetin-like 2 perform fundamental but distinct roles in the regulation of human cell migration. Fidgetin localizes to the centrosome and normally promotes cell motility. Cells depleted of fidgetin display severe reduction in motility rates. In stark contrast, fidgetin-like 2 associates with microtubules at the cell edge and normally functions to suppress cell movement. Cells lacking fidgetin-like 2 display a several fold increase in their rate of movement. Moreover, we have found that depletion of fidgetin-like 2 promotes wound healing and neovascularization in animal models. We will pursue the following two specific aims that, together, test the central hypothesis that fidgetin and fidgetin-like 2 recognize and modify distinct microtubule subpopulations thereby controlling different parameters of cell movement: Aim 1: Test the hypothesis that Fidgetin normally promotes cell motility by selectively severing and releasing microtubule minus-ends from centrosomes. Aim 2: Test the hypothesis that Fidgetin-like 2 normally suppresses cell motility by shearing the plus-ends of dynamic microtubules positioned at the cell edge. Our research plan combines complementary state-of-the-art biophysical and cellular approaches to systematically determine 1) how fidgetin and fidgetin-like 2 catalyze the removal of tubulin from the microtubule lattice, 2) how these activities are harnessed in cells to model MT arrays, and 3) how the modeling of cellular MTs by fidgetin or fidgetin-like 2 is translated into altered cell motility. Work will be carried out under the co-direction of David Sharp, an expert cell and molecular biologist, and Jennifer Ross, an expert single molecule biophysicist. Successful completion of the proposed work will provide fundamental insights into the basic mechanisms of microtubule regulation of cell motility, which is a central process in human development and health, and also foundationally establish a body of knowledge for the potential development of novel therapeutic paradigms to enhance tissue regeneration and repair through the manipulation of cell movement.

描述(由申请人提供)：微管细胞骨架的快速重塑对于正常的细胞分裂、运动和形态发生是必不可少的。参与这种重塑的一类独特而耐人寻味的蛋白质是微管切断酶，之所以被命名，是因为它们在体外能够在微管晶格中产生内部断裂。这项研究计划中的研究将阐明微管切断酶亚家族中仍鲜为人知的成员的细胞功能和生物物理作用机制，称为fidgetins。这个亚家族的创始成员fidgetin长期以来一直被认为对哺乳动物的发育很重要，但其发育功能的机制基础仍然不清楚。在最近的一项研究中，我们发现人的fidgetin是一种微管切断酶和负端解聚酶。我们现在发现fidgetin和与fidgetin密切相关的蛋白fidgetin-like 2在调节人类细胞迁移中发挥着基本但不同的作用。Fidgetin定位于中心体，通常促进细胞运动。缺乏fidgetin的细胞表现出运动率的严重下降。与之形成鲜明对比的是，《坐立不安2》 与细胞边缘的微管相联系，通常起到抑制细胞运动的作用。缺少fidgetin-like 2的细胞显示其运动速度增加了几倍。此外，我们还发现，在动物模型中，缺失fidgetin-like 2可以促进伤口愈合和新生血管形成。我们将追求以下两个特定的目标，共同检验中心假说，即fidgetin和fidgetin-like 2识别和修饰不同的微管亚群，从而控制不同的细胞运动参数：目标1：检验fidgetin通过选择性地切断和释放中心体上的微管负端来促进细胞运动的假说。目的2：验证Fidgetin-like 2通常通过剪切位于细胞边缘的动态微管的正端来抑制细胞运动的假设。我们的研究计划结合互补的最先进的生物物理和细胞方法来系统地确定1)fidgetin和fidgetin-like 2如何催化微管晶格中微管蛋白的去除，2)这些活动如何在细胞中被利用来模拟MT阵列，以及3)fidgetin或fidgetin-like 2对细胞MT的建模如何转化为细胞运动性的改变。这项工作将在细胞和分子生物学家大卫·夏普和单分子生物物理学家詹妮弗·罗斯的共同指导下进行。这项拟议工作的成功完成将为微管调节细胞运动的基本机制提供基本的见解，这是人类发育和健康的核心过程，并为潜在开发新的治疗范例建立基础知识体系，以通过操纵细胞运动来增强组织再生和修复。