Epigenetic Control of Heterochromatin Formation

异染色质形成的表观遗传控制

• 批准号: 6821250
• 负责人: RABINDRANATH DE LA FUENTE
• 金额: $ 28.53万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-07-01 至 2007-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6821250
• 关键词: DNA methylation; RNA interference; centromere; chromosome movement; developmental genetics; genetic regulation; genetically modified animals; helicase; heterochromatin; laboratory mouse; meiosis; nuclear proteins; nucleic acid repetitive sequence; oogenesis; pancreatic ribonuclease; plasmids; protein binding; protein localization; protein structure function; transfection /expression vector

DESCRIPTION (provided by applicant): Centromeric heterochromatin formation is essential to regulate fundamental aspects of nuclear architecture, gene silencing through DNA methylation, and chromosome segregation. The appropriate regulation of these processes is critical for the successful completion of oogenesis. Although the mechanisms involved in the establishment of heterochromatin domains during oogenesis are currently unknown, chromatin-remodeling proteins are likely to be important determinants. The focus of this proposal will be on ATRX, a member of the SWI/SNF2 family of chromatin remodeling proteins. This proposal is based on the hypotheses that: 1) chromatin remodeling proteins such as ATRX are important components of centromeric heterochromatin domains in mammalian oocytes; 2) that ATRX is essential for proper chromosome segregation during meiosis; and 3) that binding of ATRX to centromeric domains is essential for DNA methylation at satellite centromeric sequences. The specific objectives include: (1) The functional characterization of the ATRX protein in mouse oocytes; (2) To determine the mechanisms by which ATRX is bound to centromeric heterochromatin during oogenesis; (3) To determine whether ATRX is involved in establishing the patterns of DNA methylation at centromeric satellite sequences during the transition from meiosis to the first mitosis. A transgenic RNA interference (RNAi) approach will be used to generate an oocyte-specific conditional knockdown of the ATRX transcript. This mouse model will be used to establish whether functional ablation of ATRX disrupts appropriate chromosome-microtubule interactions during meiosis and the transition from gamete to embryo. Moreover, whether ATRX is involved in methylation of minor or major satellite DNA sequences will be determined. These studies will provide new information regarding the relationship between DNA methylation, heterochromatin formation and chromosome segregation during female meiosis, as well as the role of meiotic centromere function in the transmission of a euploid chromosome complement.

描述（由申请人提供）：丝粒异染色质形成对于调节核结构的基本方面，通过DNA甲基化和染色体分离至关重要。这些过程的适当调节对于成功完成卵子是至关重要的。尽管目前未知在卵子发生过程中涉及异染色质结构域涉及的机制，但染色质复制蛋白可能是重要的决定因素。该提案的重点将放在ATRX上，ATRX是染色质重塑蛋白的SWI/SNF2家族的成员。该建议基于以下假设：1）染色质重塑蛋白（例如ATRX）是哺乳动物卵母细胞中丝粒异染色质结构域的重要组成部分； 2）ATRX对于减数分裂过程中适当的染色体分离至关重要； 3）ATRX与丝粒结构域的结合对于在卫星中心序列上的DNA甲基化至关重要。特定目标包括：（1）小鼠卵母细胞中ATRX蛋白的功能表征； （2）确定在卵子发生过程中ATRX与centromeric异染色质结合的机制； （3）确定在从减数分裂到第一个有丝分裂的过渡期间，在centromeric卫星序列上建立DNA甲基化模式。 转基因RNA干扰（RNAI）方法将用于生成ATRX转录本的卵母细胞特异性敲低。该小鼠模型将用于确定ATRX的功能消融是否会破坏减数分裂过程中适当的染色体 - 微管相互作用以及从配子到胚胎的过渡。此外，将确定AT​​RX是否参与次要卫星DNA序列的甲基化。这些研究将提供有关女性减数分裂过程中DNA甲基化，异染色质形成和染色体隔离之间关系的新信息，以及减数分裂centromere功能在杜鹃花染色体补体传播中的作用。