Actomyosin in a Basal Body-Associated Fibrillar Complex

基底体相关纤维复合物中的肌动球蛋白

• 批准号: 9301904
• 负责人: Ray Gavin
• 金额: $ 25.27万
• 依托单位: CUNY Brooklyn College
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 1993
• 资助国家: 美国
• 起止时间: 1993-07-01 至 1996-12-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=9301904&HistoricalAwards=false
• 关键词: Actomyosin; Basal; Body; Associated; Fibrillar

In ciliated cells, basal bodies are contained within separate, filamentous cages connected to basal body microtubules. A recent study showed that actin is a component of the basal body cage complex in Tetrahymena and may be part of an actomyosin system associated with these organelles (Hoey and Gavin, J. Cell. Sci., 103, November 1992). There are three main objectives to this project. The first is to identify the myosin component of the putative actomyosin system in the Tetrahumena basal body cage complex; the second is to localize myosin within the complex; and the third is to determine whether basal body cage myosin mediates actin filament motility. An anti-oral apparatus anti-serum will be screened for cross-reactivity with myosins I and II on immunoblots. Antibodies that cross-react with myosin polypeptides will be affinity-purified and used to localize myosin-like components of the Tetrahymena basal body cage complex using immunogold labeling. To show actin filament interactions, in vitro assembled actin filaments will be mixed with purified basal body cage myosin and applied directly to a carbon-coated grid, negatively stained, and viewed by electron microscopy. %%% One of the most important forms of eukaryotic cellular motility is that mediated by axonemal beating: i.e., the highly regulated back- and-forth movement of slender, microtubule-containing cellular extensions (cilia or flagella). Depending on circumstances, such movements enable a cell to either swim in a regulated manner or to create local currents in its vicinity resulting in the movement of the immediate environment with respect to the cell. Specific cases of ciliary or flagellar motility include such diverse examples as: swimming of mammalian sperm toward the ovum, a process critical to mammalian reproduction; swimming of unicellular algae or protozoans in pondwater; feeding of protozoans by sweepinging food-containing pondwater into their oral cavities; and clearance of respiratory mucus from the lungs by the ciliary beat of epithelial cells lining the trachea and bronchi. The axonemes of such cells are rooted in cellular structures called basal bodies, which are homologous to centrosomes, structures in which intracellular microtubular arrays (such as mitotic spindles) are rooted. Although basal bodies have been studied for more than a century, important questions about their structure, chemistry, formation, and function remain unanswered. These organelles form definite structural associations with various fibrillar complexes in the cell, one of which is the cage. The structure-function relationship of these fibrillar complexes has not received much attention from investigators. Recent work has shown actin to be present in the basal body cage complex of a ciliated protozoan, Tetrahymena (a useful model system for studying ciliary function), and on the basis of the presence of actin it was proposed that there is an actomyosin contractile machinery involved in the regulation of basal body position (and by implication, the direction of the movement). This proposal focuses on the identification of the putative myosin component. The implications of such a model go well beyond motility in protozoans.

在纤毛细胞中，基体包含在单独的， 连接到基体微管的丝状笼。 最近的一 研究表明，肌动蛋白是基体笼的组成部分， 一个复杂的四膜虫，可能是肌动球蛋白系统的一部分 与这些细胞器相关（Hoey和Gavin，J. Cell.科学， 103，November 1992）. 这有三个主要目标 项目 首先是鉴定肌球蛋白成分， Tetrahumena基体笼中的推定肌动球蛋白系统 第二是将肌球蛋白定位在复合物内;以及 第三是确定基底体笼肌球蛋白是否介导 肌动蛋白丝运动性 一种抗口腔器械抗血清将 在免疫印迹上筛选与肌球蛋白I和II的交叉反应性。 与肌球蛋白多肽交叉反应的抗体将被免疫。 亲和纯化并用于定位肌球蛋白样成分， 四膜虫基体笼复合体的免疫金标记。 为了显示肌动蛋白丝的相互作用， 细丝将与纯化的基体笼肌球蛋白混合， 直接应用于碳涂层网格，负染色，和 通过电子显微镜观察。 %%% 真核细胞运动的最重要形式之一是 由轴丝搏动介导的：即，受到高度监管的背部- 细长的、含微管的细胞 延伸（纤毛或鞭毛）。 视情况而定，如 运动使细胞能够以有规律的方式游动， 在其附近产生局部电流， 与细胞相关的直接环境。 具体案件 纤毛或鞭毛运动的例子包括： 哺乳动物精子向卵子游动，这是一个关键过程， 哺乳动物的繁殖;单细胞藻类或原生动物的游动 在池塘水中;通过清扫含食物的食物来喂养原生动物 池塘水进入他们的口腔;和清除呼吸道 粘液由肺上皮细胞的纤毛搏动衬里 气管和支气管 这些细胞的轴丝扎根于 细胞结构称为基体，这是同源， 中心体，细胞内微管阵列的结构 (such有丝分裂纺锤体）生根。 尽管基底体具有 已经研究了世纪，重要的问题是， 它们的结构、化学性质、形成和功能 无人回应 这些细胞器形成明确的结构协会 细胞中有各种纤维状复合物，其中之一是 笼子 这些原纤维的结构与功能关系 复合物没有受到研究人员的太多关注。 最近的工作表明肌动蛋白存在于基体笼 复合体的纤毛虫原生动物，四膜虫（一个有用的模式系统 用于研究睫状体功能），并且基于 肌动蛋白，有人提出，有一个肌动球蛋白收缩 参与调节基础身体位置的机械（以及 （即运动的方向）。 该提案重点 对肌球蛋白成分的鉴定 的 这种模型的意义远远超出了原生动物的能动性。