CHROMOSOME ARCHITECTURE AND DOMAINS OF REPRESSION

染色体结构和抑制域

• 批准号: 2906766
• 负责人: Marc R. Gartenberg
• 金额: $ 31.64万
• 依托单位: UNIV OF MED/DENT NJ-R W JOHNSON MED SCH
• 依托单位国家: 美国
• 财政年份: 1994
• 资助国家: 美国
• 起止时间: 1994-09-12 至 2003-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2906766
• 关键词: DNA replication; Saccharomyces cerevisiae; chromatin; chromosomes; fungal genetics; gene induction /repression; genetic manipulation; genetic recombination; immunoprecipitation; nucleic acid sequence

The long-range goal of this project is to determine how large chromosomal domains are transcriptionally inactivated by packaging DNA into heterochromatin. This form of heritable repression inhibits some genes permanently and others in a developmentally regulated manner. Chromosomal translocations that move genes between transcriptionally- poised domains and heterochromatic domains lead to aberrant expression states and diseases. Leukemias and other cancer frequently arise from the inappropriate expression of translocated genes. Genom rearrangements that position a gene near heterochromatin result in an epigenetic pattern of expression: the gene is "off" in some cells and "on" in others and these expression states are maintained stably across many generations. This bimodal pattern of expression is attributed to the clonal propagation of heterochromatin structure that has spread along DNA to varying extents in different cells. In yeast Saccharomyces cerevisiae, genes near telomeres and the silent mating-type loci are repressed by a structure, termed silent chromatin, that bears remarkable similarity to heterochromatin. At the silent mating loci, repression requires cis-actin regulatory sequences, termed silencers, and a set of non-histone chromatin components known as the Sir proteins. To identify basic principles underling heterochromatin- like repression, a host of novel approaches will be used to investigate the structure, stability, and inheritance of silent chromatin in yeast. A primary strategy will involve the use of inducible site-specific recombination to form DNA rings of silent chromatin in vivo. Biochemically isolated rings will be used to analyze the structure and composition of silent chromatin and perform functional studies of transcriptional repression. To investigate silent chromatin stability, the persistence of transcriptional repression in rings uncoupled from chromosomal silencers will be analyzed in vivo. The heritable propagation of silent chromatin will be investigated by analyzing the establishment of the repressive structure on newly replicated templates. Factors that can function as boundaries of silent chromosomal domains will be identify by a simple genetic selection.

该项目的远距离目标是确定通过将DNA包装到异染色质中将大型染色体域被转录灭活。这种可遗传的抑制形式以发育监管的方式永久抑制了一些基因，而另一些基因。在转录结构域和异质域之间移动基因的染色体易位会导致异常的表达状态和疾病。白血病和其他癌症经常来自易位基因的不当表达。将基因在异染色质附近定位的基因重排会导致表达的表观遗传模式：在某些细胞中，基因在其他细胞中“关闭”，而在其他细胞中则“ ON”，并且这些表达态在许多世代中保持稳定。这种表达的双峰模式归因于异染色质结构的克隆传播，该结构已沿着DNA扩散到不同细胞中的变化扩展。在酿酒酵母的酵母菌糖疗中，端粒附近的基因和沉默的交配型基因座被一种称为无声染色质的结构所抑制，与异染色质具有显着相似之处。在沉默的交配基因座，抑制需要顺式 - 肌动蛋白调节序列，称为消音器，以及一组被称为siR蛋白的非内酮染色质成分。为了确定构成异染色质抑制作用的基本原理，将使用许多新颖的方法来研究酵母中无声染色质的结构，稳定性和遗传。主要策略将涉及使用诱导位点特异性重组以形成体内静音染色质的DNA环。生化分离的环将用于分析无声染色质的结构和组成，并对转录抑制进行功能研究。为了研究无声的染色质稳定性，将在体内分析与染色体消音器未偶联的环中转录抑制的持续性。无声染色质的遗传传播将通过分析新复制模板上的压制结构的建立来研究。可以通过简单的遗传选择来确定可以用作无声染色体结构域边界的因素。