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: 7659636
• 负责人: ROBERT A MARTIENSSEN
• 金额: $ 41.2万
• 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 2011-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7659636
• 关键词: Animals; Archaea; Arts; Binding Proteins; Cataloging; Catalogs; Centromere; Chromatin; Chromosomes; Cleaved cell; Complex; DNA Damage; DNA Polymerase II; DNA Transposable Elements; Epigenetic Process; Evolution; Fission Yeast; Gene Expression Regulation; Genes; Genetic Screening; Genetic Transcription; Genome; Heterochromatin; Histone H3; Histones; Imagery; In Vitro; Junk DNA; Lighting; Lysine; Mammals; Mediating; Methylation; Methyltransferase; MicroRNAs; Microscopy; Morphology; Nuclear Envelope; Nuclear Pore; Pathway interactions; Plants; Polyadenylation; Process; Proteins; RNA; RNA Interference; Recruitment Activity; Reporter Genes; Resolution; Role; Site; Slice; Specificity; Structure; System; Techniques; Tertiary Protein Structure; Testing; Transcript; Transposase; Visual; base; cohesin; demethylation; gene therapy; genetic analysis; histone modification; human disease; in vivo; insight; mRNA Cleavage and Polyadenylation Factors; mutant; novel; positional cloning; promoter; ultra high resolution

DESCRIPTION (provided by applicant): The pericentromeric repeats of the fission yeast Schizosaccharomyces pombe are transcribed and subject to RNA interference (RNAi). This influences heterochromatic histone modification, which silences reporter genes and recruits proteins required for centromere function, including cohesin. This mechanism is conserved in mammals and plants, and has implications both for gene therapy as well as for the epigenetic basis for human disease. In fission yeast, the Rik1 complex is related to both cleavage and polyadenylation specificity factor (CPSF) and to DNA damage binding protein (DDB1) complexes and is required for RNAi and histone modification. It is recruited to heterochromatin by association with the highly conserved, transposase-related protein Cenp-B, in a process that also requires cleavage (slicing) of nascent heterochromatic transcripts by Argonaute. We will investigate how transcripts are initiated by Pol II and processed by RNAi and the exosome. We will determine how Rik1 is recruited by RNAi, how it influences histone methylation and demethylation, and how it interacts with the nuclear envelope. We will use state-of- the-art microarray profiling techniques, as well as ultra high resolution Structured Illumination microscopy. S. pombe is a particularly useful system to study RNAi mediated heterochromatin, because this seems to be the only function of RNAi in fission yeast (there are no known miRNA) and mutants are generally viable. It should be possible therefore to take one or more of these screens to saturation, providing a unique catalog of genes involved in the primitive function of RNAi. This "function appears to be conserved in archaebacteria, fission yeast, protozoans, plants and animals, and may provide insight into the evolution of the chromosome, and its relationship to repetitive or "junk" DNA. Transposable elements and other repeats are common to all of these genomes, and may hold the key to epigenetic mechanisms of gene regulation and chromosome function.

描述（由申请人提供）：裂殖酵母粟酒裂殖酵母的着丝粒周围重复序列被转录并受到RNA干扰（RNAi）。这会影响异染色质组蛋白修饰，从而沉默报告基因并招募着丝粒功能所需的蛋白质，包括粘连蛋白。这种机制在哺乳动物和植物中是保守的，对于基因治疗以及人类疾病的表观遗传基础都有影响。在裂殖酵母中，Rik1 复合物与切割和多聚腺苷酸化特异性因子 (CPSF) 以及 DNA 损伤结合蛋白 (DDB1) 复合物相关，并且是 RNAi 和组蛋白修饰所必需的。它通过与高度保守的转座酶相关蛋白 Cenp-B 结合而被招募到异染色质，这一过程还需要 Argonaute 对新生异染色质转录物进行切割（切片）。我们将研究转录如何由 Pol II 启动并由 RNAi 和外泌体处理。我们将确定 Rik1 如何被 RNAi 招募、它如何影响组蛋白甲基化和去甲基化，以及它如何与核膜相互作用。我们将使用最先进的微阵列分析技术以及超高分辨率结构照明显微镜。粟酒裂殖酵母是研究 RNAi 介导的异染色质特别有用的系统，因为这似乎是裂殖酵母中 RNAi 的唯一功能（没有已知的 miRNA），并且突变体通常是可行的。因此，应该有可能使这些筛选中的一个或多个达到饱和，从而提供参与 RNAi 原始功能的独特基因目录。这种“功能似乎在古细菌、裂殖酵母、原生动物、植物和动物中是保守的，并且可以提供对染色体进化及其与重复或“垃圾”DNA 的关系的深入了解。转座元件和其他重复是所有生物所共有的这些基因组的组成部分，可能掌握着基因调控和染色体功能的表观遗传机制的关键。