Novel CpG-free vertebrate insulator: role for YY1

新型无 CpG 脊椎动物绝缘体：YY1 的作用

• 批准号: 8113359
• 负责人: PRABHAKARA P REDDI
• 金额: $ 7.39万
• 依托单位: UNIVERSITY OF VIRGINIA
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-08-01 至 2013-01-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8113359
• 关键词: Address; Beckwith-Wiedemann Syndrome; Binding Proteins; Binding Sites; Biological Assay; CCCTC-binding factor; Cell Culture Techniques; Cell Cycle Proteins; Chickens; Chromatin; Chromatin Loop; Code; Colorectal Cancer; Complex; Consensus; Couples; DNA; DNA Methylation; DNA Sequence; DNA-Binding Proteins; Data; Developmental Process; Drosophila genus; EMSA; Elements; Enhancers; Environment; Excision; Failure; Feasibility Studies; Fertility; Future; Gene Expression Regulation; Genes; Genetic Transcription; Genome; Genomic Imprinting; Germ Cells; Germ Lines; Globin; Heterochromatin; Human; Human Genome; In Vitro; Incidence; Indium; Infertility; Knockout Mice; Lead; Longitudinal Studies; Male Infertility; Mammals; Mediating; Meiosis; Methylation; Modeling; Mus; Muscular Dystrophies; Mutation; Nuclear Lamina; Nuclear Matrix; Physiological; Pilot Projects; Play; Production; Protein Binding; Proteins; RNA Interference; Reporting; Research Project Grants; Retinoblastoma; Role; Signal Transduction; Site; Specificity; Sperm-Ovum Interactions; Spermatids; Spermatogenesis; Structure; Testing; Testis; Tissues; Transcriptional Silencer Elements; Transgenic Mice; United States; Vertebrates; Work; X Inactivation; Yang; acrosomal protein SP-10; age group; base; beta Globin; cofactor; gene repression; human disease; imprint; in vivo; male; men; mouse model; novel; prevent; promoter; protein TDP-43; public health relevance; reproductive; sperm cell; transcription factor

DESCRIPTION (provided by applicant): Insulators are regulatory DNA sequences which form gene boundaries and protect genes from the influence of neighboring chromatin. Insulator-proteins bind to insulator DNA elements in the genome and tether those regions to subnuclear structures thus forming chromatin loops such that enhancer/silencer elements of one gene may not gain access to the promoter of a neighboring gene. Given the complexity of the human genome, one would expect that a variety of insulators and insulator- proteins regulate transcription of the estimated 35,000 genes. For example, in Drosophila alone, four types of insulators and binding proteins have been reported. In contrast, only one type of insulator (chicken globin HS4) and insulator-protein (CTCF) has been reported so far in vertebrates. CTCF mediated insulator activity is modulated by methylation of DNA. We have characterized a CpG-free, CTCF-independent vertebrate insulator (SP-10 insulator) from a testis-specific gene. The SP-10 insulator is conserved between mice and men. Preliminary mutation studies have indicated that transcription factor YY1 may mediate the SP-10 insulator function. In the proposed pilot study, we will investigate if YY1 interacts with the SP-10 insulator in vitro (using EMSA) and in a physiological context (using ChIP) (Specific Aim 1); and address the requirement of YY1 for SP-10 insulator function using RNAi in a cell culture model as well as a YY1 conditional knockout mouse model (Specific Aim 2). This feasibility study is a small self-contained research project with the potential to lead to a larger hypothesis-driven study. A recent study has shown that YY1 is essential for spermatogenesis. The current study will expand the role of YY1 to testis-specific gene transcription and in the long-term to male infertility. PUBLIC HEALTH RELEVANCE: Incidence of infertility is on the rise; it has been estimated that in the United States one in six couples in the reproductive age group are unable to conceive. The male factor contributes to 50% of the infertility cases. Treatment options are limited for men owing to a poor understanding of the mechanisms controlling sperm production (spermatogenesis). Here we propose to study the role of a transcription factor YY1 in regulating testis-specific gene transcription. Since the mouse model has recently shown that absence of YY1 in male germ cells leads to defective spermatogenesis, the proposed studies on the function of YY1 in testis are highly important for male infertility and are therefore of high public health relevance.

描述（申请人提供）：绝缘子是形成基因边界并保护基因免受相邻染色质影响的调控DNA序列。绝缘子蛋白结合到基因组中的绝缘子DNA元件上，并将这些区域拴在亚核结构上，从而形成染色质环，使得一个基因的增强子/沉默子元件不能接近相邻基因的启动子。考虑到人类基因组的复杂性，人们会认为各种各样的绝缘子和绝缘子蛋白调控着大约35,000个基因的转录。例如，仅在果蝇中，已经报道了四种类型的绝缘体和结合蛋白。与此相反，迄今为止在脊椎动物中仅报道了一种类型的绝缘子（鸡珠蛋白HS 4）和绝缘子蛋白（CTCF）。CTCF介导的绝缘子活性由DNA甲基化调节。我们已经从睾丸特异性基因中鉴定了一种无CpG、CTCF独立的脊椎动物绝缘子（SP-10绝缘子）。SP-10绝缘子在小鼠和人之间是保守的。初步的突变研究表明，转录因子YY 1可能介导SP-10绝缘子功能。在拟议的初步研究中，我们将研究YY 1是否在体外（使用EMSA）和生理环境（使用ChIP）中与SP-10绝缘子相互作用（具体目标1）;并在细胞培养模型以及YY 1条件性敲除小鼠模型中使用RNAi解决YY 1对SP-10绝缘子功能的需求（具体目标2）。这项可行性研究是一个小型的独立研究项目，有可能导致更大的假设驱动的研究。最近的一项研究表明，YY 1是精子发生所必需的。目前的研究将YY 1的作用扩展到睾丸特异性基因转录和长期男性不育。 公共卫生相关性：不孕症的发病率正在上升;据估计，在美国，每六对育龄夫妇中就有一对无法怀孕。男性因素导致50%的不孕症病例。由于对控制精子产生（精子发生）的机制缺乏了解，男性的治疗选择有限。在这里，我们建议研究转录因子YY 1在调节睾丸特异性基因转录中的作用。由于最近的小鼠模型表明，YY 1在男性生殖细胞中的缺失导致精子发生缺陷，因此建议对YY 1在睾丸中的功能进行研究对于男性不育症非常重要，因此具有高度的公共卫生相关性。