The Biochemical Basis for the Mechanics of Cytokinesis

描述（由申请人提供）：胞质分裂是细胞形状变化的一个引人注目的例子，在此期间，收缩环的机械收缩导致有丝分裂结束时的细胞分离。胞质分裂对于正常细胞增殖是必需的，并且由于其在过度增殖性疾病如癌症中的作用而具有医学意义。 在盘基网柄藻中，已经确定了两种具有互补细胞分布的肌动蛋白交联蛋白之间的遗传相互作用。皮质素-I定位于收缩环，而动力皮质素是皮质富集的，但被排除在收缩环之外。这意味着细胞已经进化出不同的肌动蛋白交联蛋白，其具有互补的细胞分布，协调细胞形状的变化，并且这些不同的肌动蛋白交联蛋白可以控制区域皮质粘弹性。 在这个提议中，为了确定dynacortin如何控制粘弹性，将使用纯化的蛋白质和各种平衡和动力学技术研究其肌动蛋白交联机制。将使用多种体内试验研究动力皮质素的细胞作用，包括野生型细胞中的显性效应、皮质素-I的抑制和动力皮质素功能丧失突变体的拯救。由于胞质分裂是一个机械过程和肌动蛋白细胞骨架是主要贡献者细胞的粘弹性，我们假设，dynacortin和cortexillin-I控制区域粘弹性。我们正在使用激光跟踪显微流变学来测量界面的粘弹性模量，并划分野生型和基因工程菌株，其中dynacortin，cortexillin-I和其他活动已被改变。事实上，在初步实验中，dynacortin和cortexillin-I是皮质粘弹性的重要调节剂。 新的基因参与皮质形状控制将被确定使用的遗传抑制皮质素-I和肌球蛋白-II的遗传增强。肌球蛋白-II是位于收缩环的主要机械力发生器。一种新的蛋白质，DdERM，这被确定为一个cortexillin-I抑制剂，假设系皮质肌动蛋白的质膜，并有助于皮质粘弹性。DdERM是两类哺乳动物肌动蛋白相关蛋白的融合体，即ezrin-radixin-moesin（ERM）和fimmectin。因此，这种分子是相当大的兴趣，因为它在皮质功能和细胞形状的控制和其不寻常的域结构的作用。

DESCRIPTION (provided by applicant): Cytokinesis is a dramatic example of a cell shape change during which the mechanical constriction of the contractile ring leads to cell separation at the end of mitosis. Cytokinesis is essential for normal cell proliferation and is of medical interest for its role in hyperproliferative diseases such as cancer. In Dictyostelium discoideum, genetic interactions have been identified between two actin cross-linking proteins that have complementary cellular distributions. Cortexillin-I is localized to the contractile ring while dynacortin is cortically enriched but excluded from the contractile ring. The implication is that cells have evolved distinct actin cross-linking proteins with complementary cellular distributions that orchestrate cell shape changes, and these different actin cross-linking proteins may control regional cortical viscoelasticity. In this proposal, to ascertain how dynacortin controls viscoelasticity, its actin cross-linking mechanism will be studied using purified proteins and a variety of equilibrium and kinetic techniques. The cellular role of dynacortin will be studied using a variety of in vivo assays including dominant effects in wild type cells, suppression of cortexillin-I and rescue of a dynacortin loss-of-function mutant. Because cytokinesis is a mechanical process and the actin cytoskeleton is the principal contributor to the cell's viscoelasticity, we hypothesize that dynacortin and cortexillin-I control regional viscoelasticity. We are using laser-tracking microrheology to measure the viscoelastic moduli of interphase and dividing wild type and genetically engineered strains where dynacortin, cortexillin-I and other activities have been altered. Indeed, in preliminary experiments, dynacortin and cortexillin-I are significant modulators of cortical viscoelasticity. New genes involved in cortical shape control will be identified using genetic suppression of cortexillin-I and genetic enhancement of myosin-II. Myosin-II is the major mechanical force generator located at the contractile ring. One novel protein, DdERM, which was identified as a cortexillin-I suppressor, is hypothesized to tether the cortical actin to the plasma membrane and contribute to cortical viscoelasticity. DdERM is a fusion of two classes of mammalian actin-associated proteins, ezrin-radixin-moesin (ERM) and fimbrin. Thus, this molecule is of considerable interest because of its role in cortical function and cell shape control and its unusual domain structure.