Meiotic Centromere Behavior in Yeast

描述（由申请人提供）： 在人类中，减数分裂I中的错误是出生缺陷、智力迟钝的主要原因，也是不孕症的重要原因。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。人类的非整倍体有时是因为姐妹染色单体在减数分裂I中分离而发生的，就像它们在有丝分裂中一样。其他时候，非整倍体的发生是因为同源配偶体在减数分裂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分离控制的减数分裂特异性机制实现的。首先，减数分裂I分离是通过重组将同源染色体系在一起来完成的，因此它们将充当分离伴侣。其次，着丝粒被改变，保护着丝粒粘连蛋白不被去除，所以姐妹染色单体保持连接。第三，一个以前未被认识到的过程，同源着丝粒配对（CEN配对），以帮助它们在纺锤体上定位的方式将着丝粒保持在一起。第四，纺锤体的组装被延迟，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。任何这些减数分裂对染色体行为的修饰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这些减数分裂特定的过程和地址：如何减数分裂I着丝粒组装，使姐妹染色单体将分离作为一个单位？CEN配对的基础是什么？纺锤体的组装如何与染色体的行为协调，以使减数分裂着丝粒的重组得以正确完成？该提案分为三组实验。这些实验使用芽殖酵母作为模式生物，并结合了分子遗传学、细胞成像和生物化学方法。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，并将它们替换为决定减数分裂I分离行为的动粒。这些实验将采用质谱方法，这将允许评估整体动粒组成的细胞通过减数分裂I的进展，和互补的成像方法，将检查个别动粒组件的行为。第二组实验将研究如何完成CEN配对。CEN配对需要联会复合体（SC）蛋白，Zip 1，它仍然存在于配对的着丝粒SC解体后。最后一组实验采用遗传学方法来确定染色体和纺锤体动力学在减数分裂中是如何协调的，以便纺锤体的形成被阻止，直到染色体正确识别并与它们的分离伴侣配对。完成拟议的实验将阐明如何减数分裂重构的动粒结构，以及着丝粒和纺锤体的行为，允许同源染色体在减数分裂I分离高保真。

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.