Determining how the spindle assembly checkpoint monitors chromosome biorientation

描述（由申请人提供）：染色体分离必须仔细调节，因为错误分离会导致灾难性的后果，如基因组不稳定和癌症。为了正确分离，所有复制的染色体必须在有丝分裂纺锤体上正确定向，每一对姐妹着丝点都附着在来自相反纺锤体极点的微管上。即使单个染色体无法实现这种“生物定向”配置，也会触发一个主要的细胞周期检查点反应，称为纺锤体组装检查点（SAC），它会停止细胞周期并阻止姐妹染色单体的分离。许多SAC的下游效应物已经被很好地表征，但是染色体错位如何触发SAC初始激活的机制细节仍然知之甚少。在这一调控中起关键作用的是染色体乘客复合体（CPC），它控制着必要的有丝分裂激酶Aurora b的活性

DESCRIPTION (provided by applicant): Chromosome segregation must be carefully regulated because missegregation has catastrophic consequences such as genome instability and cancer. In order to segregate properly, all replicated chromosomes must orient correctly on the mitotic spindle with each pair of sister kinetochores making attachments to microtubules from opposing spindle poles. Failure of even a single chromosome to achieve this "bioriented" configuration triggers a major cell cycle checkpoint response, known as the spindle assembly checkpoint (SAC), which halts the cell cycle and prevents separation of sister chromatids. Many of the downstream effectors of the SAC have been well characterized, but the mechanistic details of how misaligned chromosomes trigger the initial activation of the SAC remain poorly understood. A critical player in this regulation is the chromosomal passenger complex (CPC) that controls the activity of the essential mitotic kinase Aurora B. Highlighting the importance of the CPC to cell cycle control, members of CPC are known to be upregulated in cancer cells and are currently being used as targets in drug development. However, despite this importance a thorough understanding of the functions of CPC components is lacking. The research training plan proposed here describes three complementary lines of experimentation to investigate the mechanisms by which the CPC controls the activity of Aurora B and thereby coordinates chromosome structure and cell cycle progression. These studies will provide critical insights into chromosome biology and the regulation of the cell cycle that will have broad relevance to genome maintenance as well as specific impacts on the understanding of cancer progression. The goals of the research training plan proposed here are 1) to determine how the CPC effects the localization and dynamic turnover of Aurora B on mitotic chromosomes 2) to elucidate which targets of Aurora B kinase are effected by chromosome biorientation and 3) to uncover whether the interaction with specific binding partners confers the sensitivity of CPC function to chromosome structure. To meet these goals, three complementary lines of experimentation will be undertaken using the Xenopus egg extract system, which has been widely used to characterize the biochemistry of cell cycle regulation because of the unprecedented control it provides of cell cycle progression. The power of the Xenopus system will be combined with high-resolution microscopic analysis and recent innovations in the reconstitution and purification of mitotic chromatin. These experiments will test current hypotheses for the function of the CPC and will provide valuable training in the application of cutting edge technologies to the important fields of chromosome biology and cell cycle control.