Isolation and Characterization of Yeast rDNA Chromatin

酵母 rDNA 染色质的分离和表征

• 批准号: 7155837
• 负责人: ERICA Y JACOBS
• 金额: $ 4.6万
• 依托单位: ROCKEFELLER UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-09-01 至 2008-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7155837
• 关键词: Saccharomyces cerevisiae; binding sites; chimeric proteins; chromatin; electrospray ionization mass spectrometry; fungal genetics; genetic regulation; genetic transcription; green fluorescent proteins; histones; immunoprecipitation; matrix assisted laser desorption ionization; nucleic acid purification; nucleic acid repetitive sequence; postdoctoral investigator; posttranslational modifications; ribosomal DNA; transcription factor

DESCRIPTION (provided by applicant): How do genes turn on and off? If DNA sequence remains unchanged, changes in transcription are caused by differences in DNA-associated factors. So understanding chromatin composition changes within particular regions of DNA is vital to understanding gene expression. This proposal aims to better understand the role of chromatin dynamics in gene expression by examining how ribosomal DNA (rDNA) chromatin composition relates to rDNA transcription in the yeast Saccharomyces cerevisiae using mass spectrometry. This project has three specific aims. The first is to purify native rDNA chromatin from yeast cells. Second, the identity [and locations] of bound proteins and nature of histone modifications will be determined using mass spectrometry. Third, differences in chromatin composition and histone modifications between actively transcribed and silent rDNA will be determined. Future directions include extending the technology from yeast to mammalian cells. Chromatin composition at crucial loci could be followed through developmental transudations, environmental changes, or during oncogenesis. My ultimate goal is the complete characterization of an organism's "chromatome:" the identity, quantity, and location of all proteins associated with every DNA sequence and their modification states through development, aging and disease.

描述（由申请人提供）：基因如何开启和关闭？如果 DNA 序列保持不变，则转录的变化是由 DNA 相关因子的差异引起的。因此，了解 DNA 特定区域内染色质组成的变化对于了解基因表达至关重要。该提案旨在通过使用质谱法检查核糖体 DNA (rDNA) 染色质组成与酿酒酵母中 rDNA 转录的关系，更好地了解染色质动力学在基因表达中的作用。该项目有三个具体目标。第一个是从酵母细胞中纯化天然 rDNA 染色质。其次，将使用质谱法确定结合蛋白的身份[和位置]和组蛋白修饰的性质。第三，将确定活跃转录和沉默 rDNA 之间染色质组成和组蛋白修饰的差异。未来的方向包括将该技术从酵母扩展到哺乳动物细胞。关键位点的染色质组成可以通过发育渗出、环境变化或肿瘤发生过程来追踪。我的最终目标是对生物体“染色体”的完整表征：与每个 DNA 序列及其在发育、衰老和疾病过程中的修饰状态相关的所有蛋白质的身份、数量和位置。