Meiotic Centromere Behavior in Yeast

描述(由申请人提供)： 在人类中，减数分裂I的错误是导致出生缺陷、智力低下的主要原因，也是导致不孕的重要因素。在减数分裂I中，染色体表现出独特的分离模式；姐妹染色单体保持在一起，而同源染色体彼此分离。人类非整倍体的发生有时是因为姐妹染色单体在减数分裂I中分离，就像在有丝分裂中一样。有时，非整倍体的发生是因为同源配对在减数分裂I时分离到纺锤体的同一侧。正确的减数分裂分离模式是通过改变染色体结构和细胞周期控制分离的减数分裂特有机制实现的。首先，减数分裂I的分离是通过重组将同源染色体捆绑在一起来完成的，因此它们将作为分离伙伴。其次，动粒被改变，着丝粒粘连蛋白不被移除，因此姐妹染色单体保持连接。第三，一种以前未知的过程，同源着丝粒配对(CEN配对)，将着丝粒以帮助它们在纺锤体上定向的方式结合在一起。第四，纺锤体的组装被推迟，因此减数分裂染色体对的形成可以在染色体开始分离之前完成。这些对染色体行为的减数分裂修饰中的任何一个的失败都可能是人类减数分裂分离错误的罪魁祸首。这里提出的实验集中在减数分裂的特定过程和地址：减数分裂是如何组装的，从而使姐妹染色单体作为一个单位进行分离？CEN配对的基础是什么？纺锤体组装如何与染色体行为相协调，以便正确完成减数分裂着丝粒的重组？该提案分为三组实验。这些实验使用萌芽酵母作为模式生物，并结合了分子遗传学、细胞成像和生化方法。第一组实验将探索当减数分裂开始时，细胞如何分解染色体上的动粒，并用决定减数分裂I分离行为的动粒取而代之。这些实验将使用质谱学方法，可以评估细胞在减数分裂I过程中的整体动粒组成，以及互补的成像方法，可以检查单个动粒成分的行为。第二组实验将考察CEN配对是如何完成的。CEN配对需要突触膜复合体(Synaptonemal Complex，SC)蛋白ZIP1，在SC分解后，ZIP1保持在配对的着丝粒上。最后一组实验使用遗传学方法来确定染色体和纺锤体动态在减数分裂中是如何协调的，从而阻止纺锤体的形成，直到染色体被正确识别并与它们的分离伙伴配对。拟议实验的完成将阐明减数分裂重塑着丝粒结构以及着丝粒和纺锤体行为如何允许同源染色体在减数分裂I中高保真地分离。

DESCRIPTION (provided by applicant): In humans, errors in meiosis I are the leading cause of birth defects, mental retardation, and a significant contributor to infertility. In meiosis I, chromosomes exhibit a unique segregation pattern; sister chromatids remain together, while homologous chromosomes segregate from each other. Human aneuploidy sometimes occurs because sister chromatids separate in meiosis I, as they do in mitosis. Other times aneuploidy occurs because homologous partners segregate to the same side of the spindle at meiosis I. The correct meiotic segregation pattern is achieved by meiosis-specific mechanisms that alter chromosome structures and the cell cycle control of segregation. First, meiosis I segregation is accomplished by tethering the homologous chromosomes through recombination so they will act as segregation partners. Second, kinetochores are altered, and centromeric cohesins are protected from removal, so sister chromatids remain joined. Third, a previously unrecognized process, homologous centromere pairing (CEN-pairing), holds the centomeres together in ways that help them orient on the spindle. Fourth, the assembly of the spindle is delayed so the formation of meiotic chromosome pairs can be completed before the chromosomes begin segregating. Failures in any of these meiotic modifications to chromosome behavior could be culprits in human meiotic segregation errors. The experiments proposed here focus on these meiosis-specific processes and adress: How are meiosis I kinetochores assembled so that sister chromatids will segregate as a unit? What is the basis for CEN-pairing? How is spindle assembly coordinated with chromosome behavior to allow proper completion of the re-structuring of meiotic centromeres? The proposal is divided into three sets of experiments. The experiments use budding yeast as a model organism and a combination of molecular genetic, cell imaging and biochemical approaches. The first set of experiments will explore how the cell disassembles the kineotchores that are on chromosomes when meiosis begins, and replaces them with kinetochores that will dictate meiosis I segregation behaviors. These experiments will employ mass spectrometry methods that will allow assessment of overall kinetochore composition as cells progress through meiosis I, and complementary imaging approaches that will examine behaviors of individual kinetochore components. The second set of experiments will examine how CEN-pairing is accomplished. CEN-pairing requires the synaptonemal complex (SC) protein, Zip1, which persists at the paired centromeres after SC disassembly. The final set of experiments employs genetic methods to determine how chromosome and spindle dynamics are coordinated in meiosis so that spindle formation is blocked until chromosomes have properly identified and paired with their segregation partners. Completion of the proposed experiments will elucidate how meiotic remodeling of kinetochore structures, and centromere and spindle behaviors, allows homologous chromosomes to be segregated with high fidelity in meiosis I.