The Role of Stu1 in Mitotic Spindle Stability

描述(由申请人提供)：在细胞分裂过程中遗传物质的适当分离是至关重要的，这取决于由微管和相关蛋白质组成的双极有丝分裂纺锤体的形成和稳定性。正如在癌症和遗传病中观察到的那样，错误分离导致的非整倍体细胞受到有害影响并降低了生存能力。酿酒酵母蛋白Stulp与微管相互作用，定位于有丝分裂纺锤体中区，对正确的纺锤体结构和活力是必不可少的。然而，与Stu1相互作用的蛋白质以及Stu1p在有丝分裂纺锤体的形成和维持中的作用尚不清楚。使用串联亲和纯化(TAP)和质谱学鉴定Stu1相关蛋白将有助于理解Stu1在纺锤体中的上下文：酵母是一个有价值的遗传系统，但深入的生物化学是有限的。非洲爪哇卵提取物是研究纺锤体形成和微管动力学的一个强大的体外系统。使用从编码Stu1同源物的cDNA文库中鉴定的一种非洲爪哇蛋白的抗体进行免疫定位和免疫耗竭，将允许分析这类蛋白在双极纺锤体形成和微管动力学中的作用。

DESCRIPTION (provided by applicant): Proper segregation of genetic material during cell division is critical and depends on the formation and stability of the bipolar mitotic spindle composed of microtubules and associated proteins. Aneuploid cells resulting from mis-segregation suffer detrimental effects and reduced viability as observed in cancers and genetic diseases. The Saccharomyces cerevisiae protein Stulp interacts with microtubules, localizes to the mitotic spindle midzone region, and is essential for proper spindle structure and viability. However, proteins that interact with Stu1 and the role of Stu1p in formation and maintenance of the mitotic spindle is not understood. Identification of Stu1 associating proteins using Tandem Affinity Purification (TAP) and mass spectroscopy will aid in understanding the context of Stu1 in the spindle: Yeast is a valuable genetic system, but in depth biochemistry is limited. Xenopus laevis egg extracts are a powerful in vitro system for studying spindle formation and microtubule dynamics. Immunolocalization and immunodepletion of extracts using antibodies against a Xenopus protein identified from cDNA libraries encoding a Stu1 homologue will allow analysis of this class of proteins in bipolar spindle formation and microtubule dynamics.