Genetic and Epigenetic Determinants of Centromere Identity

项目摘要 着丝粒是介导着丝粒组装和精确定位的基本染色体元件。 染色体分离。着丝粒缺陷导致染色体错误分离，带来有害影响 细胞和生物体的健康和生育能力。在大多数多细胞物种中，着丝粒由大的 染色质标记的含有着丝粒特异性组蛋白变体的高重复DNA区域 CENP-A以前的工作表明着丝粒DNA和CENP-A染色质都有可能 启动着丝粒活动从头开始；然而，它们各自对着丝粒规范的贡献 有丝分裂和减数分裂仍然难以捉摸。这项提案的总体目标是确定着丝粒 DNA和染色质有助于着丝粒的同一性。后生动物的着丝粒对 由于它们的大小和高度重复的性质，完全测序和组装阻碍了我们的能力 系统地询问着丝粒DNA元件的作用。此外，目前还没有系统 以测试缺乏着丝粒DNA的新着丝粒是否能维持着丝粒 有丝分裂和减数分裂的功能和特性。利用我们在果蝇中的独特进步， 这包括其着丝粒序列的鉴定和组装以及建立 可诱导的从头着丝粒系统，这项建议将：1)测试染色质介导的着丝粒是否能维持 通过发育和减数分裂进行染色体分离，有效取代内源着丝粒；2) 测试着丝粒DNA元件如何有助于CENP-A染色质建立的特定假设 或者是维护。总而言之，这项工作将有助于揭示果蝇的着丝粒指定机制， 一种特殊的模型系统，允许在动物发育和生育的背景下进行着丝粒研究- 与包括人类在内的其他物种具有广泛的相关性。

Project Summary Centromeres are essential chromosomal elements that mediate kinetochore assembly and accurate chromosome segregation. Centromere defects lead to chromosome missegregation, with detrimental effects on cell and organism health and fertility. In most multicellular species, centromeres are composed of large regions of highly repetitive DNA marked by chromatin containing the centromere-specific histone variant CENP-A. Previous work demonstrated that both centromeric DNA and CENP-A chromatin have the potential to initiate centromere activity de novo; however, their respective contributions to centromere specification in mitosis and meiosis have remained elusive. The overall goal of this proposal is to determine how centromeric DNA and chromatin contribute to centromere identity. The centromeres of metazoans have been refractory to full sequencing and assembly due their large size and highly repetitive nature, hampering our ability to systematically interrogate the role of centromeric DNA elements. Additionally, there are currently no systems in which to test if de novo centromeres, which are devoid of centromeric DNA, and can sustain centromere function and specification through mitotic and meiotic divisions. Using our unique advancements in Drosophila, which include the identification and assembly of its centromeric sequences and the establishment of an inducible de novo centromere system, this proposal will: 1) test if chromatin-mediated centromeres can sustain chromosome segregation through development and meiosis, effectively replacing endogenous centromeres; 2) test specific hypothesis on how centromeric DNA elements may contribute to CENP-A chromatin establishment or maintenance. Collectively, this work will shed light into centromere specification mechanisms in Drosophila, an exceptional model system that allows centromere studies in the context of animal development and fertility- with broad relevance to other species, including humans.