The Role of Retroelements in Centromere Function

逆转录元件在着丝粒功能中的作用

• 批准号: 10468779
• 负责人: Rachel O'Neill
• 金额: $ 43.91万
• 依托单位: UNIVERSITY OF CONNECTICUT STORRS
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-09-19 至 2024-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10468779
• 关键词: Address; Artificial Human Chromosomes; Biological Assay; Cell Cycle; Cell division; Cells; Centromere; Chromatin; Chromatin Modeling; Chromosomal Instability; Chromosome 17; Chromosome Segregation; Chromosome abnormality; Chromosomes; Clustered Regularly Interspaced Short Palindromic Repeats; Complex; Computing Methodologies; DNA Sequence; DNA-Directed RNA Polymerase; Data; Defect; Deposition; Elements; Engineering; Epigenetic Process; Evaluation; Event; Failure; Fostering; Genetic; Genetic Transcription; Genome; Genomic Instability; Genomics; Histones; Human; Human Chromosomes; Infertility; Kinetochores; Link; Location; Maintenance; Malignant Neoplasms; Mediating; Meiosis; Mitosis; Modeling; Molecular; Molecular Chaperones; Normal Cell; Nucleosomes; Outcome; Outcome Study; Pathway interactions; Play; Process; Production; Proteins; RNA; Retroelements; Retrotransposition; Ribonucleoproteins; Role; Satellite DNA; Satellite RNA; Site; Specificity; Technology; Testing; Time; Transcript; Transcriptional Regulation; Untranslated RNA; Variant; Work; centromere protein A; chromosome missegregation; design; experimental study; genetic element; human disease; in vivo; innovation; insight; novel strategies; promoter; recruit; spatiotemporal; telophase

The essential function of centromeres in chromosome segregation during cell division requires a complex cascade of epigenetic events involving changes to chromatin character and kinetochore assembly. Kinetochore formation during mitosis is the culmination of a cycle defined by the “loading”, or deposition, of newly synthesized CENP-A, a variant histone, into centromeric chromatin. CENP-Atis faithful assembly during late telophase/early G1 of the cell cycle is facilitated by its histone chaperone, HJURP. This assembly cascade is sensitive to perturbation by genetic, epigenetic and environmental insults, with catastrophic consequences for genome/cell stability, but the genomic elements that guide accurate CENP-A nucleosome assembly are not well understood. A central conundrum in understanding the genomic features that aQract CENP-A nucleosome assembly is the observation that established centromeres are replete with satellite DNA while de novo centromeres (e.g. neocentromeres) lack satellites, yet are defined by retroelements, such as LINE-1s. While it appears CENP-A nucleosome occupancy may not require specific DNA sequences, mounting evidence demonstrates that RNA is a critical component of the epigenetic cascade leading to faithful CENP-A nucleosome assembly. However, the sequence specificity, spatiotemporal requirements for and transcriptional regulation of these centromeric RNAs are currently unknown. In Preliminary Data, we show that centromeric retroelements (cenTEs) are sites of engaged RNA polymerase and are involved in the CENP-A assembly cascade at human centromeres, linking a common transcribed genomic feature to CENP-A assembly at both native and de novo centromeres in humans for the first time. Leveraging our expertise in centromere assembly, noncoding RNAs, chromosome engineering, and genomics, we have formulated three aims, each with an innovative approach that will allow us to provide an unbiased assessment of where within centromeres transcription initiates, when during the cell cycle transcript initiation and elongation occur, and how these transcripts mediate centromere nucleosome assembly. We will use chromosome engineering to directly test whether cenTEs and their transcriptional activity are sufficient to facilitate de novo centromere assembly on chromosomes. These engineered chromosomes provide a new model to study the processes guiding cenTE-mediated centromere assembly and stabilization, and to examine misregulated incorporation of centromeric histones. The outcomes of this study will fill a large gap in our current understanding of centromere assembly and maintenance in normal cells, and provide valuable insight into events underlying chromosome aberration presenting in human diseases, infertility, and cancers of high metastatic potential.

着丝粒在细胞分裂过程中染色体分离中的基本功能需要一个 涉及染色质特征和动粒变化的复杂级联表观遗传事件 组装件.有丝分裂过程中动粒的形成是一个由“加载”定义的循环的顶点， 或沉积，新合成的CENP-A，一种变异的组蛋白，进入着丝粒染色质。CENP-Atis 在细胞周期末期/G1期早期的忠实装配是由其组蛋白促进的 监护人HJURP这种装配级联对遗传、表观遗传和 环境损伤，对基因组/细胞稳定性造成灾难性后果，但基因组 指导CENP-A核小体准确组装的元件还没有很好地理解。中央 理解基因组特征的难题是aQract CENP-A核小体组装是 观察到已建立的着丝粒充满了卫星DNA，而从头着丝粒 (e.g. neocentromeres）缺乏卫星，但由retroelements定义，如LINE-1。虽然 CENP-A核小体占据可能不需要特定的DNA序列，越来越多的证据表明， 表明RNA是导致忠实CENP-A的表观遗传级联的关键组分。 核小体组装然而，序列特异性、时空要求和 这些着丝粒RNA的转录调控目前是未知的。在初步数据中，我们 显示着丝粒逆转录元件（cenTE）是参与RNA聚合酶的位点， 在人类着丝粒的CENP-A装配级联中，连接一个共同的转录基因组特征， 首次在人类的原生和从头着丝粒上进行CENP-A组装。利用我们 在着丝粒组装，非编码RNA，染色体工程和基因组学方面的专业知识，我们有 制定了三个目标，每个目标都有一个创新的方法，使我们能够提供一个公正的 评估着丝粒内转录起始的位置，在细胞周期中何时转录 起始和延伸发生，以及这些转录本如何介导着丝粒核小体组装。我们 将使用染色体工程直接测试cenTE及其转录活性 足以促进着丝粒在染色体上的从头组装。这些基因改造的染色体 提供了一个新的模型来研究引导cenTE介导的着丝粒组装的过程， 稳定化，并检查着丝粒组蛋白的误调节掺入。的成果 这项研究将填补我们目前对着丝粒组装和维持的理解中的一个巨大空白。 在正常细胞中，并提供有价值的洞察事件的基础染色体畸变呈现 在人类疾病、不孕症和高转移潜能的癌症中。