Meiotic Centromere Behavior in Yeast

描述(申请人提供)：在过去的30年里，文献中有零星的报道，在减数分裂的两个不同阶段，同源和非同源染色体的着丝粒之间配对。首先，对各种生物(洋葱、小麦、发芽酵母、小鼠)的研究表明，在减数分裂的早期，就在DNA复制之后，着丝粒成对排列。这种配对主要发生在非同源着丝粒之间。这种非同源的、早期的减数分裂着丝粒配对随着同源染色体在减数分裂后期前期变得对齐而溶解。其次，着丝粒也可以在减数分裂的后期主动配对。例如，对萌芽酵母和雌性果蝇的研究表明，在同源染色体完全突触的晚期前期，未能重组(非交换染色体)的配对染色体在其着丝粒(或着丝粒周围异染色质)配对。这种非交换伙伴之间的减数分裂晚期着丝粒配对促进了它们在第一次减数分裂时的分离。在发芽酵母中，着丝粒配对的早期和晚期都依赖于联会复合体(SC)蛋白ZIP1。这个项目的目的是描述减数分裂中着丝粒配对的两个阶段，测试它们是机械相关的还是不同的，并确定它们如何影响减数分裂I中染色体的分离。由于对减数分裂着丝粒配对的观察是从如此不同的实验生物体中收集的，拟议的实验可能揭示减数分裂染色体行为的一个保守的、但在很大程度上未被认识的细节。该项目被组织成五组实验。第一组实验确定与减数分裂早期着丝粒相关的ZIP1区域，并使用亲和纯化方法鉴定参与着丝粒配对早期阶段的蛋白质。关于着丝粒配对从早期到晚期的转变，人们知之甚少。第二组实验将探索这一转变，并阐明这两个阶段的关系。在最终打算配对然后分离同源染色体的过程(减数分裂I)的开始阶段，对齐非同源着丝粒的目的是什么？第三组实验验证了这样的假设，即非同源着丝粒配对阻止着丝粒附近交叉的形成。众所周知，这种交叉会破坏许多生物体的减数分裂I染色体分离的保真度。最后两组实验将首先探索ZIP1如何促进减数分裂后期前期着丝粒的配对，其次将检验这一晚期着丝粒配对促进着丝粒附着到减数分裂I纺锤体的相反两极的假设，并在这样做的过程中，在重要方面促进减数分裂I分离的保真度。 与公共卫生相关：将染色体分配给配子的过程中的错误是导致出生缺陷的主要原因。这个项目使用萌芽酵母来确定未被发现的方式，在这种方式中，染色体的一个组成部分，即着丝粒，可能有助于指导染色体运动，从而帮助防止导致出生缺陷的错误。

DESCRIPTION (provided by applicant): Over the past thirty years there have been sporadic reports in the literature of pairing between the centromeres of both homologous and non-homologous chromosomes at two different stages of meiosis. First, studies in a variety of organisms (onions, wheat, budding yeast, mice) have revealed that early in meiosis, just after DNA replication, centromeres arrange themselves in pairs. This pairing is primarily between non-homologous centromeres. This non-homologous, early, meiotic centromere pairing dissolves as homologous chromosomes become aligned in later meiotic prophase. Second, centromeres can also actively pair at a later stage of meiosis. For example, studies with budding yeast and female Drosophila have shown that partner chromosomes that have failed to recombine (non-exchange chromosomes) pair at their centromeres (or pericentric heterochromatin) in late prophase, when homologous chromosomes are completely synapsed. This late meiotic centromere pairing between non-exchange partners promotes their disjunction at the first meiotic division. In budding yeast, both the early and late stages of centromere pairing depend upon the synaptonemal complex (SC) protein, Zip1. This project aims to characterize the two phases of centromere pairing in meiosis, test whether they are mechanistically related, or distinct, and determine how they impact the segregation of chromosomes in meiosis I. Because observations of meiotic centromere pairing have been gathered from such a diverse range of experimental organisms, the proposed experiments may reveal the details of a conserved, but largely unrecognized, aspect of meiotic chromosome behavior. The project is organized into five groups of experiments. The first group of experiments determines the region of Zip1 that is involved in association with the centromere in early meiosis and uses affinity purification methods to identify proteins that are involved in the early phase of centromere pairing. Little is known about the transition from early to later centromere pairing. A second group of experiments will explore this transition and shed light on the relationship of the two stages. What is the purpose of aligning non-homologous centromeres at the beginning of a process (meiosis I) that is ultimately intended to pair then separate homologous chromosomes? A third group of experiments tests the hypothesis that non-homologous centromere pairing blocks the formation of crossovers near to centromeres. Such crossovers are known to disrupt meiosis I chromosome segregation fidelity in many organisms. The final two groups of experiments will explore first, how Zip1 promotes the pairing of centromeres in later meiotic prophase, and second, will test the hypothesis that this late centromere pairing promotes the attachment of centromeres to opposite poles of the meiosis I spindle, and in doing so, contributing to meiosis I segregation fidelity in important ways. PUBLIC HEALTH RELEVANCE: Errors in the process of distributing chromosomes to the gametes is the major cause of birth defects. This project uses budding yeast to identify undiscovered ways in which a component of the chromosome, called the centromere, might help direct chromosomes movements, in doing so helping to prevent the errors that lead to birth defects.