Novel CpG-free vertebrate insulator: role for YY1

新型无 CpG 脊椎动物绝缘体：YY1 的作用

基本信息

批准号：
7991095
负责人：
PRABHAKARA P REDDI
金额：
$ 7.7万
依托单位：
UNIVERSITY OF VIRGINIA
依托单位国家：
美国
项目类别：
财政年份：
2010
资助国家：
美国
起止时间：
2010-08-01 至 2012-07-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7991095
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 TP53 gene Tars 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 public health relevance reproductive sperm cell transcription factor

项目摘要

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（由申请人提供）：绝缘子是形成基因边界并保护基因免受邻近染色质影响的调节DNA序列。绝缘子蛋白与基因组中的绝缘子DNA元件结合，并将这些区域束缚在亚核结构上，从而形成染色质环，使得一个基因的增强子/沉默子元件可能无法接近邻近基因的启动子。鉴于人类基因组的复杂性，人们预计有多种绝缘体和绝缘体蛋白调节大约 35,000 个基因的转录。例如，仅在果蝇中，就报道了四种类型的绝缘体和结合蛋白。相比之下，迄今为止，脊椎动物中只报道了一种类型的绝缘体（鸡珠蛋白 HS4）和绝缘体蛋白（CTCF）。 CTCF 介导的绝缘体活性受 DNA 甲基化调节。我们从睾丸特异性基因中鉴定出一种无 CpG、不依赖 CTCF 的脊椎动物绝缘体（SP-10 绝缘体）。 SP-10 绝缘体在小鼠和人类之间是保守的。初步突变研究表明转录因子YY1可能介导SP-10绝缘子功能。在拟议的试点研究中，我们将研究 YY1 是否在体外（使用 EMSA）和生理环境（使用 ChIP）中与 SP-10 绝缘体相互作用（具体目标 1）；并在细胞培养模型以及 YY1 条件敲除小鼠模型中使用 RNAi 解决 YY1 对 SP-10 绝缘体功能的要求（具体目标 2）。该可行性研究是一个小型独立研究项目，有可能引发更大规模的假设驱动研究。最近的一项研究表明，YY1 对于精子发生至关重要。目前的研究将扩大 YY1 在睾丸特异性基因转录中的作用，并从长远来看对男性不育症的作用。公共卫生相关性：不孕不育的发生率正在上升；据估计，在美国，六分之一的育龄夫妇无法怀孕。 50%的不孕不育病例是由男性因素造成的。由于对控制精子产生（精子发生）的机制了解甚少，男性的治疗选择有限。在这里，我们建议研究转录因子 YY1 在调节睾丸特异性基因转录中的作用。由于小鼠模型最近表明雄性生殖细胞中缺乏 YY1 会导致精子发生缺陷，因此关于睾丸中 YY1 功能的研究对于男性不育非常重要，因此具有很高的公共卫生相关性。