MOLECULAR BIOLOGY OF MICROTUBULE-INTERACTING PROTEINS

我们的长期目标是鉴定和表征蛋白质， 组成微管并与微管相互作用。 微管对于 细胞分裂、细胞结构和细胞器在细胞中的运动。 它们也是医学上重要的抗真菌药和驱虫药的靶点 药物和癌症化疗药物。 尽管重要的是 微管，许多蛋白质是必不可少的功能， 我们的目标是确定其中的一些 proteins. 我们的总体策略是首先通过以下方法来鉴定这些蛋白质： 识别编码它们的基因，然后用这些基因来识别 并对蛋白质进行表征并确定其功能。 我们有 发现了一种这样的蛋白质，γ微管蛋白，我们现在希望测试我们的 gmma微管蛋白作为微管成核剂假说 组装件. 我们将使用丝状真菌，构巢曲霉， 研究，因为它是一个很好的有机体进行分子 基因和生物化学实验。 具体而言，我们将1） 产生γ微管蛋白基因的新的条件致死突变等位基因， 并确定这些突变对微管组装的影响， 体外和体内，2）确定γ微管蛋白抗体或突变 将阻止微管组织中心的微管组装， 3)纯化和表征γ微管蛋白。 为了识别其他基因 所需的微管功能，我们建议使用的程序， 将允许我们克隆编码蛋白质的基因， 微管，微管组织的组成部分蛋白质 中心和与γ微管蛋白相关或相互作用的蛋白。

Our long term objective is to identify and characterize proteins that compose and interact with microtubules. Microtubules are essential for cell division, cell structure, and movement of organelles through cells. They are also targets for medically important antifungal and anthelmintic agents and for cancer chemotherapy agents. In spite of the importance of microtubules, many of the proteins that are essential for the functioning of microtubules are unknown and it is our goal to identify some of these proteins. Our general strategy is to identify such proteins by first identifying the genes that encode them and then using the genes to identify and characterize the proteins and determine their functions. We have discovered one such protein, gamma tubulin, and we now wish to test our hypothesis that gmma tubulin functions as a nucleator of microtubule assembly. We will use tbe filamentous fungus, Aspergillus nidulans for our studies because it is an excellent organism for carrying out the molecular genetic and biochemical experiments we propose. Specifically we will 1) create new conditionally lethal mutant alleles of the gamma tubulin gene and determine the effects of these mutations on microtubule assembly in vitro and in vivo, 2) determine if gamma tubulin antibodies or mutations will block assembly of microtubules from microtubule organizing centers and 3) purify and characterize gamma tubulin. To identify additional genes required for microtubule functioning, we propose to use procedures that will allow us to clone genes that encode proteins that co-purify with microtubules, proteins that are components of the microtubule organizing center and proteins that are related to, or interact with, gamma tubulin.