Molecular Dissection of Cytokinesis

描述（由申请人提供）：我们试图了解动物细胞中细胞分裂，胞质分裂的分子基础。在后期纺锤体中的一组反平行的成束微管，调节以肌动球蛋白为基础的收缩环的形成，并且是完成胞质分裂所必需的。我们提出，中央纺锤体的组织和功能是一个结果的结构组织和生物化学性质的centralspindlin复杂。Centralspindlin是一种进化上保守的多聚体复合物，含有驱动蛋白样蛋白（ZEN-4/MKLP 1）和Rho家族GAP（CYK-4/MgcRacGAP），分别在后期和末期高度集中在中央纺锤体和中间体上。我们建议联合收割机在体外生物化学和在C.本研究旨在通过对线虫胚胎和哺乳动物细胞的研究，实现以下三个具体目标：（1）分析中枢纺锤蛋白复合物的分子结构，并对非典型驱动蛋白ZEN-4进行表征。通过表征centralspindlin复合物中相对孤立的关键蛋白质结构域，我们将开发必要的生化框架，以了解这些分子在更复杂的反应以及体内的功能。(2)去破译纺锤体组装的分子机制。将在体外从纯化的组分重构中心纺锤体组装，以定义这组动态组分组装成高度有序和稳定结构的原理。(3)确定中央纺锤体是如何介导胞质分裂的完成的。Centralspindlin包含两个不同的蛋白质结构域，它们被认为控制胞质分裂的后期步骤。因此，将建立这些结构域的分子功能。这项研究将提供对细胞增殖至关重要的细胞结构的分子见解。因此，我们的研究可能有助于开发用于治疗癌症的新型抗有丝分裂剂。此外，ZEN-4的分子解剖将增强我们对微管马达作用机制的理解。

DESCRIPTION (provided by applicant): We seek to understand the molecular basis of cell division, cytokinesis, in animal cells. The central spindle, a set of antiparallel bundled microtubules in the anaphase spindle, regulates formation of the actomyosin- based contractile ring and is essential for completion of cytokinesis. We propose that the organization and function of the central spindle is a consequence of the structural organization and biochemical properties of the centralspindlin complex. Centralspindlin is an evolutionary conserved, multimeric complex containing a kinesin-like protein (ZEN-4/MKLP1) and a Rho family GAP (CYK-4/MgcRacGAP) that is highly concentrated on the central spindle and midbody during anaphase and telophase, respectively. We propose to combine in vitro biochemistry and in vivo rescue assays in C. elegans embryos and in mammalian cells to address three specific aims: (1) To dissect the molecular organization of the centralspindlin complex and to characterize the atypical kinesin protein, ZEN-4. By characterizing the critical protein domains in the centralspindlin complex in relative isolation, we will develop the biochemical framework necessary to understand the function of these molecules in more complex reactions as well as in vivo. (2) To decipher the molecular mechanism of central spindle assembly. Central spindle assembly will be reconstituted in vitro from purified components in order to define the principles by which this set of dynamic components assemble into a highly ordered and stable structure. (3) To determine how the central spindle mediates completion of cytokinesis. Centralspindlin contains two distinct protein domains that are proposed to control late steps in cytokinesis. Therefore, the molecular function of these domains will be established. This research will provide molecular insights into a cellular structure that is critical for cell multiplication. Therefore our research could contribute to the development of novel anti-mitotic agents for the treatment of cancer. Furthermore, molecular dissection of ZEN-4 will enhance our understanding of the mechanism of action of microtubule motors.