Chromatin Structure during Spermatogenesis

描述（由申请人提供）：生理上和基因上完整的性染色体的精确减数分裂分离对雄性生殖和物种生存至关重要；染色体分离发生在减数分裂前期。破坏前期染色体动力学的突变导致不育和减数分裂受阻，无法修复DNA损伤。在精母细胞细胞核中，性染色体位于与常染色体不同的核区域。在常染色体常染色质和转录活跃区域，常染色体配对、突触和同源重组确保它们在减数分裂中准确分离。然而，非同源的X和Y染色体在很大程度上是不配对的，形成了一个独特的异色和转录不活跃的核区域，称为XY小体，这是本提案的重点。导致XY染色质修饰失败的突变使雄性不育；因此，适当的XY染色质动力学对于精子的成功发生至关重要。指导所提出的实验的基本假设是，未配对的染色体形成一组独特的蛋白质和翻译后蛋白质修饰，共同重塑染色质。这一假设的两个关键组成部分将被测试：异色化是否是任何未配对DNA的特性，以及特定蛋白质及其修饰在XY体中的性质和作用。首先，有人提出，像X和Y染色体一样，任何未配对的染色体都会被表观遗传修饰和转录沉默，这是一种被称为“未配对DNA减数分裂沉默”（MSUD）的一般减数分裂机制。这将在Aim 1中通过确定未配对的三体常染色体是否被修饰和沉默来测试。这一目标的结果也将深入了解修饰的染色质在诱导基因组损伤修复中的功能，这一过程对于产生具有完整染色体的配子是绝对必要的。其次，有人提出对未配对的XY染色质的识别通过募集独特的蛋白质和翻译后蛋白质修饰导致异色化。这一假设将在Aim 2中通过关注SUMO（小泛素相关修饰剂）对XY体蛋白的修饰来验证。鉴定底物蛋白之后，将进行基因破坏，以提供其在维持XY体中的功能的基因测试。这些目标的结果将促进我们对基于减数分裂染色质的机制的认识，这些机制决定了细胞核功能并确保精子发生的遗传保真度。

DESCRIPTION (provided by applicant): Accurate meiotic segregation of physically and genetically intact sex chromosomes is essential for male reproduction and species survival; chromosome segregation is set up in meiotic prophase. Mutations impairing prophase chromosome dynamics lead to sterility and arrested meiosis with failure to repair DNA damage. In the spermatocyte nucleus, sex chromosomes lie in a nuclear territory distinct from that of autosomal chromosomes. In the euchromatic and transcriptionally active domain, autosomal chromosomes pair, synapse and undergo the homologous recombination that ensures their accurate segregation in the meiotic divisions. However, the non-homologous X and Y chromosomes are largely unpaired and form a unique heterochromatic and transcriptionally inactive nuclear territory, known as the XY body, which is the focus of this proposal. Mutations causing failure in modification of XY chromatin render males sterile; therefore, appropriate XY chromatin dynamics are essential for successful spermatogenesis. The underlying hypothesis guiding the proposed experiments is that unpaired chromosomes nucleate a unique array of proteins and post-translational protein modifications that together remodel chromatin. Two (2) critical components of this hypothesis will be tested: whether heterochromatization is a property of any unpaired DNA and the nature and role in the XY body of specific proteins and their modifications. First, it is proposed, like the X and Y chromosomes, any unpaired chromosome will be epigenetically modified and transcriptionally silenced by a general meiotic mechanism known as "meiotic silencing by unpaired DNA" (MSUD). This will be tested in Aim 1 by determining if an unpaired trisomic autosomal chromosome is modified and silenced. Results from this aim will also yield insight into function of the modified chromatin in repair of induced genomic damage, a process that is absolutely essential for producing gametes with intact chromosomes. Second, it is proposed that recognition of unpaired XY chromatin leads to heterochromatization through recruitment of unique proteins and post-translational protein modifications. This hypothesis will be tested in Aim 2 by focusing on modification of XY body proteins by SUMO (small ubiquitin-related modifier). Identification of the substrate proteins will be followed by gene disruption to provide a genetic test of their function in maintenance of the XY body. Results from these aims will advance our knowledge about the meiotic chromatin-based mechanisms that determine nuclear function and ensure genetic fidelity of spermatogenesis.