Self-organization of the radial microtubule array

描述（由申请人提供）：细胞质的组织是至关重要的 对活细胞中发生的所有过程都很重要，包括 区室化、细胞代谢和细胞分裂。据信 微管系统在组织细胞质中起着关键作用 通过形成支持膜交通并确定 细胞器的位置。径向阵列的形成通常被假定为 可能是MT在中心体成核的结果，中心体通常位于 细胞的中心。然而，最近的研究表明，细胞具有 显著的自组织能力和MT电机，通常使用MT 作为“轨道”，可以依次将MT布置成径向阵列。的机制 自我组织在很大程度上仍然是未知的。拟议的总体目标 研究的目的是阐明定义形成的自组织机制 以及在体内对径向MT阵列进行定心。我们最近的观察 建立了实验系统，可以解决这些问题。的 系统涉及色素细胞，黑色素细胞的细胞质片段。在 在黑色素细胞中，成千上万的色素颗粒迅速聚集到中心， 重新分散在整个细胞质中。颗粒通过MT电机移动。 缺乏中心体的黑色素细胞的胞质片段迅速形成， 将细胞质MT的径向阵列定位到中心。形成这种 阵列需要存在色素颗粒、MT动力学和 MT马达的活性，细胞质动力蛋白。结合实验性 包括数字荧光显微镜，光漂白， 光活化、激光显微手术和运动特异性探针的显微注射 将被用来阐明自组织和自我中心的机制， 黑色素细胞碎片具体而言，我们将解开MT的机制 控制MT的空间组织的颜料聚集体上的成核。 我们将确定中心力产生的位置， MT电机在径向MT阵列定位中的作用。最后我们将 确定最低限度的补充分子组成部分的必要性， 自我组织和自我中心，通过重构这些现象， 体外这些研究对于理解 细胞组织，也用于设计新的治疗策略 旨在预防和治疗疾病。

DESCRIPTION (provided by applicant): Organization of cytoplasm is crucially important to all the processes that occur in living cells, including compartmentalization, cell metabolism, and cell division. It is believed that the system of microtubules (MTs) plays a key role in organizing the cytoplasm by forming a radial array that supports membrane traffic and determines positions of organelles. Formation of the radial array is generally assumed to be a result of MT nucleation at the centrosome, which often locates in the center of a cell. Recent studies indicate, however, that cells have a remarkable self-organizing capacity and that MT motors, which normally use MTs as 'rails', may in turn arrange MTs into a radial array. The mechanisms of self-organization remain largely unknown. The overall goal of the proposed research is to elucidate the self-organization mechanisms that define formation and centering of the radial MT array in vivo. Our recent observations established experimental system, which allows addressing these questions. The system involves cytoplasmic fragments of pigment cells, melanophores. In melanophores, thousands of pigment granules rapidly aggregate to the center or redisperse throughout the cytoplasm. The granules move by means of MT motors. Cytoplasmic fragments of melanophores lacking the centrosome rapidly form and position to the center the radial array of cytoplasmic MTs. Formation of such an array requires the presence of pigment granules, MT dynamics and the activity of a MT motor, cytoplasmic dynein. A combination of experimental techniques that include digital fluorescence microscopy, photobleaching, photoactivation, laser microsurgery and microinjection of motor-specific probes will be used to elucidate mechanisms of self-organization and self-centering in melanophore fragments. Specifically, we will unravel the mechanism of MT nucleation on the pigment aggregates that controls spatial organization of MTs. We will locate the sites of production of the centering force and determine the role for MT motors in positioning of the radial MT array. Finally, we will determine the minimal complement of molecular components essential for the self-organization and self-centering by reconstituting these phenomena in vitro. These studies are important for understanding the general mechanisms of cellular organization but also for the design of new therapeutic strategies aimed at prevention and treatment of disease.