Transcriptional Networks Regulating Luminal Environment in the Epididymis

调节附睾管腔环境的转录网络

• 批准号: 8700439
• 负责人: ANN HARRIS
• 金额: $ 28.58万
• 依托单位: LURIE CHILDREN'S HOSPITAL OF CHICAGO
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-08-10 至 2016-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8700439
• 关键词: Address; Androgen Receptor; Bioinformatics; Biological Assay; Cells; Chloride Channels; Chromatin; Chromatin Structure; Chromosomes; Complex; Control Locus; Cyclic AMP; Cystic Fibrosis; Cystic Fibrosis Transmembrane Conductance Regulator; DNA-Binding Proteins; Data; Defect; Deoxyribonuclease I; Deoxyribonucleases; Development; Disease; Duct (organ) structure; Ejaculation; Elements; Enhancers; Environment; Epididymis; Epithelial Cells; Epithelium; Fertility; Gene Expression; Gene Expression Regulation; Genes; Genital system; Genomics; Goals; Hereditary Disease; Human; In Vitro; Inborn Genetic Diseases; Infertility; Inherited; Insulator Elements; Investigation; Ion Channel; Ion Transport; Ions; Left; Link; Male Genital Organs; Male Infertility; Maps; Mediating; Methods; Modification; Molecular; Molecular Conformation; Mutate; Mutation; Obstruction; Organ; Pathway interactions; Play; Process; Proteins; Recruitment Activity; Regulation; Regulator Genes; Regulatory Element; Reporter Genes; Research; Role; Series; Site; Small Interfering RNA; Sperm Maturation; Structure; Swimming; System; Techniques; Testis; Therapeutic Intervention; Tissues; Trans-Activators; Tube; Vas deferens structure; Water; base; birth control; cell type; chromatin immunoprecipitation; cis acting element; cohesin; genome-wide; in vivo; male; malformation; novel; reproductive; research study; solute; sperm cell; three dimensional structure; vector; zygote

DESCRIPTION (provided by applicant): Functional inactivation (by mutation) of the cystic fibrosis transmembrane conductance regulator (CFTR), a small conductance, cAMP activated chloride ion channel, leads to developmental defects that cause male infertility in humans. The mechanism by which inactivation of the CFTR gene causes infertility is poorly understood but is believed to be due in part to malformations or pathological changes in the male genital duct system. The goals of the proposed research are to decipher the transcriptional networks that regulate luminal environment in the epididymis, by building on our current understanding of the mechanisms regulating expression of the CFTR gene. Mutations in CFTR cause the devastating inherited disorder cystic fibrosis (CF). The CFTR channel makes an essential contribution to ion transport and the luminal environment in the epididymis and its loss causes obstruction and/or loss of the male genital ducts and subsequent infertility. The epididymis forms the proximal part of the excretory duct system of the male reproductive tract and is a complex organ that secretes and absorbs ions, water and inorganic solutes. Critically, it provides the correct luminal environment for normal sperm maturation. In normal epididymis the luminal environment is established and maintained through the functional cooperation of multiple ion channels, ion exchangers, solute carriers and transporters, though the mechanisms that coordinate the expression of these genes has not been well studied. In the first specific aim we will determine how modifications in the chromatin structure of the CFTR locus control its expression in the epididymis. We will evaluate the cis-acting enhancers and insulator elements that likely establish the looped conformation of the active locus and determine how trans-acting factors such as CTCF and cohesin influence this spatial organization. Moreover, we will investigate mechanisms and DNA- binding proteins that may suppress gene expression in cells where CFTR is less active or inactive. Experiments in the second specific aim will determine the transcriptional pathways that coordinate expression of ion channels, ion exchangers, transporters and solute carriers that interact with CFTR in the epididymis epithelium to establish and maintain the luminal environment. We will use a genome-wide approach (DNase- seq) to identify the cis-acting regulatory elements for genes encoding the proteins that contribute to ion and solute transport across the epididymal epithelium. Following mapping of these elements we will use combined bioinformatic and experimental approaches to identify the trans-acting factors that mediate the coordinated regulation of gene expression. Successful conduct of these studies will provide unique translational opportunities to modify male fertility. In disease states we may be able to reinstate the normal epididymal luminal environment and thereby restore fertility. Through modulation of this environment we may also be able to inhibit normal sperm maturation and thereby develop novel methods of birth control.

描述（由申请人提供）：囊性纤维化跨膜电导调节器（CFTR）是一种小电导、cAMP 激活的氯离子通道，其功能失活（通过突变）会导致发育缺陷，从而导致人类男性不育。 CFTR 基因失活导致不育的机制尚不清楚，但据信部分原因是男性生殖管系统的畸形或病理变化。本研究的目标是基于我们目前对 CFTR 基因表达调节机制的理解，破译调节附睾管腔环境的转录网络。 CFTR 突变会导致破坏性遗传性疾病囊性纤维化 (CF)。 CFTR 通道对附睾中的离子传输和管腔环境做出了重要贡献，其损失会导致男性生殖管阻塞和/或损失以及随后的不育。附睾形成男性生殖道排泄管系统的近端部分，是一个分泌和吸收离子、水和无机溶质的复杂器官。至关重要的是，它为精子的正常成熟提供了正确的管腔环境。在正常附睾中，管腔环境是通过多个离子通道、离子交换剂、溶质载体和转运蛋白的功能协作来建立和维持的，尽管协调这些基因表达的机制尚未得到很好的研究。在第一个具体目标中，我们将确定 CFTR 位点染色质结构的修饰如何控制其在附睾中的表达。我们将评估可能建立活性位点环状构象的顺式作用增强子和绝缘子元件，并确定 CTCF 和粘连蛋白等反式作用因子如何影响这种空间组织。此外，我们将研究可能抑制 CFTR 活性较低或不活跃的细胞中基因表达的机制和 DNA 结合蛋白。第二个具体目标的实验将确定协调离子通道、离子交换剂、转运蛋白和溶质载体表达的转录途径，这些转录途径与附睾上皮中的 CFTR 相互作用，以建立和维持管腔环境。我们将使用全基因组方法 (DNase-seq) 来识别编码蛋白质的基因的顺式作用调控元件，这些蛋白质有助于离子和溶质跨附睾上皮的转运。在绘制这些元件后，我们将使用生物信息学和实验相结合的方法来识别介导基因表达协调调节的反式作用因子。这些研究的成功进行将为改变男性生育能力提供独特的转化机会。在疾病状态下，我们也许能够恢复正常的附睾管腔环境，从而恢复生育能力。通过调节这种环境，我们或许还能够抑制正常精子的成熟，从而开发出新的节育方法。