THE FORMATION AND FUNCTION OF MICROTUBULES

微管的形成和功能

• 批准号: 3270111
• 负责人: RAYMOND E STEPHENS
• 金额: $ 9.79万
• 依托单位: MARINE BIOLOGICAL LABORATORY
• 依托单位国家: 美国
• 财政年份: 1978
• 资助国家: 美国
• 起止时间: 1978-09-01 至 1988-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3270111
• 关键词: adenosinetriphosphatase; cell differentiation; cell membrane; chemical structure function; cilium; egg /ovum; electrofocusing; electron microscopy; flagellum; freeze etching; gel electrophoresis; genetic regulation; nonmammalian vertebrate embryology; radiotracer; saltwater environment; thermodynamics; tissue /cell culture

The mechanism of microtubule-related motility is being approached through a detailed study of the chemistry of tubulin and its interactions with itself and other proteins. We ultimately hope to be able to define the dynamics of tubulin synthesis, storage, mobilization, organelle-specific utilization, and recycling in order to better understand the role of microtubules in both cell division and cell differentiation. Four major projects will be emphasized in this continuing study. We will extend our differential detergent binding methodology for tubulin variant separation through: a) high-resolution acid-urea-Triton electrophoretic separation; b) isoelectric focusing in the presence of certain anionic, cationic, and zwitterionic detergents; c) charge-shift electrophoresis of dimeric tubulin; and d) chromatographic resolution of tubulins in accord with relative hydrophobicity. How many tubulin variants occur in different microtubules, are some shared, and what is their fate in development? We will continue primary structural studies of organelle-specific tubulines through the separation and analysis of unique peptide sequences obtained by enzymatic or chemical cleavage of tubulin subunits. What kinds of sequence variation are involved in organelle-specificity and how do these correlate with microtubule properties? We will resume our investigation of basal body and basal rootlet biochemistry through: a) characterization of tubulin solubilized from the isolated molluscan basal apparatus, comparing it with ciliary axonemal tubulins; b) characterization of rootlet protein with respect to self-assembly, interaction with basal microtubules, and the effect of calcium ions on rootlet length and periodicity; and c) reconstitution of outer doublet microtubules onto basal body templates. What is the role of the basal apparatus in ciliary assembly and function? We will explore the disposition of membrane tubulin in natural and reconstituted ciliary membranes through: a) vectorial labeling and specific cleavage to map polypeptide topology; b) incorporation of tubulin fragments into lipid bilayers; c) determination of specific lipids associated with membrane tubulin; and d) interaction of tubulin-containing membranes with cytoplasmic proteins. How is tubulin associated with the membrane, topologically, and how does it interact with cellular constituents?

微管相关运动的机制正在通过 详细研究微管蛋白的化学性质及其与自身的相互作用 和其他蛋白质。 我们最终希望能够定义动态 微管蛋白合成、储存、动员、细胞器特异性 的利用和回收，以便更好地了解其作用 微管参与细胞分裂和细胞分化。 四大 这项持续研究将重点关注项目。 我们将延长我们的 用于微管蛋白变体分离的差异去污剂结合方法 通过：a)高分辨率的酸-尿素-Triton电泳分离； b) 在某些阴离子、阳离子和离子存在下进行等电聚焦 两性离子洗涤剂； c) 二聚体的电荷转移电泳 微管蛋白； d) 微管蛋白的色谱分离度符合 相对疏水性。 不同的微管蛋白有多少种变体 微管中有些是共有的，它们在发育过程中的命运如何？ 我们 将继续细胞器特异性微管的初级结构研究 通过分离和分析获得的独特肽序列 微管蛋白亚基的酶促或化学裂解。 有哪几种顺序 变异涉及细胞器特异性以及它们如何相关 具有微管特性？ 我们将恢复对基础的调查 身体和基础支根的生物化学通过：a）表征 从分离的软体动物基底装置中溶解的微管蛋白，比较 它与睫状轴丝微管蛋白； b) 根蛋白的表征 关于自组装、与基底微管的相互作用以及 钙离子对细根长度和周期性的影响；和c) 将外部双联体微管重建到基底体模板上。 基底器在纤毛组装和功能中起什么作用？ 我们将探索膜微管蛋白在自然和环境中的分布。 通过以下方式重建睫状膜：a）矢量标记和 特异性切割以绘制多肽拓扑图； b) 微管蛋白的掺入 碎片形成脂质双层； c) 特定脂质的测定 与膜微管蛋白相关； d) 含微管蛋白的相互作用 膜上有细胞质蛋白。 微管蛋白如何与 膜的拓扑结构以及它如何与细胞相互作用 选民？