Molecular Analysis of Pericentric Sister Chromatid Cohesion

中心周围姐妹染色单体凝聚力的分子分析

• 批准号: 8005517
• 负责人: PAUL Connor MEGEE
• 金额: $ 29.8万
• 依托单位: UNIVERSITY OF COLORADO DENVER
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-07-01 至 2013-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8005517
• 关键词: Address; Anaphase; Biochemical Genetics; Biological; Biological Models; Biology; Cell Cycle; Cell division; Cell physiology; Cells; Centromere; Chromatin; Chromatin Loop; Chromosome Arm; Chromosome Segregation; Chromosome Structures; Chromosomes; Complex; Congenital Disorders; DNA; DNA Double Strand Break; DNA Repair; DNA Repair Gene; DNA Replication Timing; DNA-Directed RNA Polymerase; Data; Deposition; Development; Dimensions; Distal; Dosage Compensation (Genetics); Down Syndrome; Elements; Epigenetic Process; Eukaryota; Event; Gene Expression Regulation; Genes; Genetic; Genetic Transcription; Genomics; Goals; Growth; Hereditary Disease; Human; Intercistronic Region; Intergenic Sequence; Kinetochores; Lead; Location; Maintenance; Malignant Neoplasms; Mediating; Metabolism; Modeling; Molecular; Molecular Analysis; Morphogenesis; Nature; Nuclear; Pathway interactions; Play; Polymerase; Positioning Attribute; Process; Proteins; RNA Polymerase II; Recruitment Activity; Research; Role; Saccharomyces cerevisiae; Saccharomycetales; Shapes; Sister; Sister Chromatid; Spatial Distribution; Spontaneous abortion; Testing; Vertebrates; X Inactivation; Yeasts; arm; base; centromere protein A; centromere protein C; chromatin modification; chromatin remodeling; cohesin; cohesion; daughter cell; design; insight; interest; mutant; novel; protein complex; public health relevance; research study; segregation; transcription termination; transmission process; tumorigenesis

DESCRIPTION (provided by applicant): Accurate chromosome segregation is essential for the successful transmission of genetic information to daughter cells, and deficiencies in this process are associated with miscarriages, congenital disorders, and tumorigenesis. Integral to proper chromosome segregation is the cohesion or physical association of replicated sister chromatids, which is mediated by the evolutionarily conserved cohesin complex. Cohesins are involved not only in chromosome segregation, but also play important roles in DNA repair and gene regulation. Cohesin's spatial distribution on budding yeast chromosomes is highly reproducible, suggesting that it plays important roles in chromosome structure and function. Cohesins are frequently found in intergenic regions between convergently transcribed genes, indicating interplay between cohesin distributions and transcription that is currently not characterized. Notably, extensive cohesin-enriched domains assemble in pericentromeric (kinetochore-flanking) regions, where they promote chromosome biorientation and resist precocious sister chromatid separation. We showed previously that budding yeast kinetochores direct pericentromeric cohesin domain assembly. Our preliminary data indicate that pericentromeric cohesin domains are assembled epigenetically by a nucleation and spreading mechanism. We test the veracity of this model in Specific Aim 1 by determining whether loop formation adjacent to the centromere impedes cohesin recruitment in distal sequences. Pericentromeric chromatin will also be isolated and its protein composition characterized in an unbiased screen for kinetochore-associated factors or epigenetic chromatin modifications that direct cohesin domain assembly. Lastly, insulators that limit cohesin recruitment will be used to delineate the minimally effective pericentromeric domain necessary for high fidelity chromosome transmission. Preliminary data also indicate that cohesin distributions are directed by the prior association of the Scc2/Scc4 cohesin loader with intergenic sequences. In the second aim, we endeavor to determine the mechanisms and significance of loader and cohesin localization on chromosome arms. The order and interdependence of association of the RSC ATP-dependent chromatin remodeler, Scc2/Scc4, and cohesin will be determined at chromosome arm cohesin-associated regions to dissect the pathway for cohesin deposition. The contribution of RNA polymerase II-dependent transcription in the establishment of cohesin loader distributions will be examined following polymerase inactivation and the mechanism and significance of cohesin redistribution in an RNA polymerase II transcription termination mutant are examined to delineate the nature of the relationship between transcription and cohesin localization. The role of budding yeast cohesins in the promotion of RNA polymerase II transcription termination will be determined using conditional cohesin mutants. PUBLIC HEALTH RELEVANCE: Chromosomes must be accurately duplicated and segregated to daughter cells during each cell division, and errors in these events can lead to cancer or genetic disease, such as Down syndrome. The accurate segregation of replicated chromosomes, or sister chromatids, to daughter cells requires that sisters physically associate with one another throughout much of the cell cycle. In this application, we endeavor to understand how the proteins that mediate the physical association of sister chromatids, called cohesins, are recruited to proper locations on the chromosome to mediate chromosome segregation, DNA repair, and gene regulation, each of which is important for the maintenance of genomic integrity.

描述(申请人提供)：准确的染色体分离对于成功地将遗传信息传递到子代细胞是至关重要的，而这一过程中的缺陷与流产、先天性疾病和肿瘤的发生有关。适当的染色体分离所必需的是复制的姐妹染色单体之间的凝聚或物理联系，这是由进化上保守的粘附素复合体介导的。粘附素不仅参与染色体分离，而且在DNA修复和基因调控中发挥重要作用。粘附素在发芽酵母染色体上的空间分布具有很高的重复性，表明它在染色体结构和功能中起着重要的作用。粘附素经常发现于可转换转录基因之间的基因间隔区，这表明粘附素分布与转录之间的相互作用目前尚不清楚。值得注意的是，广泛的粘附素富集区聚集在着丝粒周围(动粒侧翼)区域，在那里它们促进了染色体的双向定位，并抵制了早熟姐妹染色单体的分离。我们以前已经证明，发芽酵母动点直接引导着丝粒周围粘连蛋白结构域的组装。我们的初步数据表明，着丝粒周围的粘连蛋白结构域是通过成核和扩散机制在表观遗传上组装的。我们通过确定着丝粒附近的环形成是否阻碍远端序列中的粘附素招募来测试该模型在特定目标1中的准确性。还将分离着丝粒周围染色质，其蛋白质组成的特征是无偏筛选动粒相关因子或指导粘附素结构域组装的表观遗传染色质修饰。最后，限制粘附素募集的绝缘体将被用来描绘高保真染色体传递所必需的最低有效的着丝粒周围区域。初步数据还表明，粘附素的分布是由Scc2/Scc4粘附素加载器与基因间序列的先前关联决定的。在第二个目标中，我们努力确定加载器和粘附素在染色体臂上定位的机制和意义。依赖于RSC ATP的染色质重构体、Scc2/Scc4和粘附素的关联顺序和相互依赖关系将在染色体臂粘附素相关区域确定，以剖析粘附素沉积的途径。在聚合酶失活后，将检测依赖于RNA聚合酶II的转录在粘附素载入器分布建立中的贡献，并研究RNA聚合酶II转录终止突变体中粘附素重新分布的机制和意义，以描述转录和粘附素定位之间关系的本质。发芽酵母粘附素在促进RNA聚合酶II转录终止中的作用将使用条件粘附素突变体来确定。 与公共卫生相关：在每次细胞分裂过程中，染色体必须准确复制并分离到子代细胞，这些事件中的错误可能会导致癌症或遗传性疾病，如唐氏综合症。复制的染色体或姐妹染色单体准确地分离到子代细胞，需要姐妹在细胞周期的大部分时间里相互联系。在这一应用中，我们努力了解调节姐妹染色单体物理联系的蛋白质，称为粘附素，是如何被招募到染色体上的适当位置来调节染色体分离、DNA修复和基因调节的，其中每一项都对维持基因组的完整性很重要。