RNAi, Histone Modification and the DDB1/CPSF-Like Complex Rik1

RNAi、组蛋白修饰和 DDB1/CPSF 样复合物 Rik1

• 批准号: 8239195
• 负责人: ROBERT A MARTIENSSEN
• 金额: $ 37.47万
• 依托单位: COLD SPRING HARBOR LABORATORY
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-08-01 至 2015-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8239195
• 关键词: Aging; Binding Proteins; Biological Models; Cell Cycle; Cell division; Centromere; Childhood; Chromosome Structures; Chromosomes; Complex; DNA; DNA Damage; DNA Polymerase II; DNA Repair; DNA Sequence; DNA biosynthesis; DNA-Directed DNA Polymerase; DNA-Directed RNA Polymerase; Disease; Ensure; Enzymes; Epigenetic Process; Failure; Fission Yeast; Functional RNA; Gene Cluster; Gene Expression; Gene Silencing; Genes; Genetic; Genetic Recombination; Genetic Transcription; Genome; Health; Heterochromatin; Histones; Human; Hybrids; Inherited; Malignant Neoplasms; Mammals; Mediating; Mental Retardation; Mitosis; Modeling; Modification; Nature; Nucleotide Excision Repair; Phase; Plants; Positioning Attribute; Proteins; RNA; RNA Interference; RNA replication; Recruitment Activity; Repetitive Sequence; Replication Origin; Research; Role; S Phase; Small RNA; Specificity; Testing; Time; Transcription-Coupled Repair; Transfer RNA; X Inactivation; Yeasts; chromatin modification; chromosome replication; cullin 4A; gene function; genetic analysis; histone modification; homologous recombination; imprint; mutant; paralogous gene; prevent; protein B; recombinational repair; repaired; transcription factor TFIIH; ubiquitin ligase; ubiquitin-protein ligase

DESCRIPTION (provided by applicant): Heterochromatin comprises tightly compacted repetitive regions of eukaryotic chromosomes. It is inherited through mitosis and has roles in transcriptional silencing, centromere specification and genome integrity, which profoundly impact epigenetic mechanisms in human health and disease. We have found that the epigenetic inheritance of heterochromatin in fission yeast requires RNA interference (RNAi) to guide histone modification, which occurs during the DNA replication phase of the cell cycle. In the fission yeast S.pombe centromeric repeats have an alternating arrangement of small RNA clusters and origins of replication that makes collision of the transcription and replication machineries all but inevitable. We propose that RNA interference promotes release of RNA polymerase (PolII) during S phase, allowing completion of centromeric DNA replication by the leading strand DNA polymerase. DNA Polymerase epsilon directly recruits the histone-modifying Rik1 complex and so can spread heterochromatin along with DNA replication. In the absence of RNAi, stalled forks are repaired by homologous recombination (HR) without histone modification, so that HR is essential in the absence of RNAi. This model may explain the participation of non-coding RNA and DNA replication in many examples of epigenetic silencing, including paramutation in plants, and imprinting and X-inactivation in mammals. S.pombe is an outstanding model system for cell cycle research, heterochromatic silencing, and RNAi. We will examine the roles of DNA replication, RNA Polymerase release, DNA recombination and repair in heterochromatic histone modification mediated by RNAi. We will utilize models of heterochromatic nucleation and RNAi, as well as chromosome profiling and genetic analysis, to test our hypothesis. We will build on our recent results concerning the roles of the Rik1 complex and Centromere-binding protein B in DNA replication and repair, as well as RNA interference. PUBLIC HEALTH RELEVANCE: Epigenetic mechanisms alter gene function independent of DNA sequence, and have profound effects on health and disease. RNA interference impacts these mechanisms by guiding the modification of histones associated with the DNA, ensuring specificity and avoiding inappropriate gene silencing. We have found that replication of the chromosome during cell division occurs at the same time as RNA interference, and that these mechanisms interact to cause silencing. The key molecules involved are conserved from yeast to humans, and are implicated in childhood disease, mental retardation, aging and cancer. Our findings suggest that therapies that target these molecules may also impact gene expression and chromosome organization. We will investigate the underlying mechanism to determine the causes and consequences of RNAi- mediated modification of chromatin during the DNA replication phase of the cell cycle.

描述(申请人提供)：异染色质由真核细胞染色体紧密致密的重复区域组成。它通过有丝分裂遗传，在转录沉默、着丝粒规范和基因组完整性方面发挥作用，从而深刻影响人类健康和疾病的表观遗传机制。我们发现，分裂酵母中异染色质的表观遗传需要RNA干扰(RNAi)来指导组蛋白修饰，组蛋白修饰发生在细胞周期的DNA复制阶段。在分裂酵母中，S.pombe着丝粒重复序列具有小RNA簇和复制起点的交替排列，这使得转录和复制机制几乎不可避免地发生碰撞。我们认为，在S期，RNA干扰促进了RNA聚合酶的释放，使领先的链DNA聚合酶完成着丝粒复制。DNA聚合酶epsilon直接招募组蛋白修饰的Rik1复合体，因此可以在DNA复制的同时传播异染色质。在没有RNAi的情况下，停滞的叉子可以通过同源重组(HR)修复，而不需要组蛋白修饰，所以在没有RNAi的情况下，同源重组是必不可少的。这个模型可以解释许多表观遗传沉默的例子中非编码RNA和DNA复制的参与，包括植物中的参数突变，以及哺乳动物中的印记和X失活。S.pombe是研究细胞周期、异染色质沉默和RNAi的优秀模型系统。我们将研究DNA复制、RNA聚合酶释放、DNA重组和修复在RNAi介导组蛋白异染色质修饰中的作用。我们将利用异染色质成核和RNAi模型，以及染色体图谱和遗传分析来检验我们的假设。我们将基于我们最近的结果，即Rik1复合体和着丝粒结合蛋白B在DNA复制和修复以及RNA干扰中的作用。 公共卫生相关性：表观遗传机制改变独立于DNA序列的基因功能，并对健康和疾病产生深远影响。RNA干扰通过引导与DNA相关的组蛋白的修饰、确保特异性和避免不适当的基因沉默来影响这些机制。我们发现，在细胞分裂过程中，染色体的复制与RNA干扰同时发生，这些机制相互作用，导致沉默。涉及的关键分子从酵母到人类都是保守的，与儿童疾病、智力低下、衰老和癌症有关。我们的发现表明，针对这些分子的治疗也可能影响基因表达和染色体组织。我们将研究潜在的机制，以确定RNAi介导的染色质修饰在细胞周期的DNA复制阶段的原因和后果。