RHOX action in Sertoli development and function

RHOX 在支持细胞发育和功能中的作用

基本信息

批准号：
9766336
负责人：
JAMES Arthur MACLEAN
金额：
$ 24.49万
依托单位：
SOUTHERN ILLINOIS UNIVERSITY CARBONDALE
依托单位国家：
美国
项目类别：
财政年份：
2018
资助国家：
美国
起止时间：
2018-08-17 至 2020-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9766336
关键词：
Ablation Address Adult Affect Androgens Antibodies Apoptosis Birth Blood-Testis Barrier CRISPR/Cas technology Carrying Capacities Cell Count Cell Differentiation process Cell Survival Cells ChIP-seq Couples Data Defect Development Down-Regulation Embryo Event Exhibits Family Female Fertility Genes Genetic Genetic Transcription Germ Cells Goals Gonadal Ridge Gonadal structure Health Homeobox Homeobox Genes Homeodomain Proteins Human Infertility Knock-out Knockout Mice Knowledge Light Link Mammals Meiosis Modeling Mus Mutation Nurses Oligospermia Output Ovulation Pathway interactions Phenotype Population Positioning Attribute Production Puberty Publishing RNA Interference Reproduction Reproductive History Research Role Sexual Development Signal Transduction Somatic Cell Specific qualifier value Sperm Count Procedure Sperm Motility Spermatogenesis Structure of primordial sex cell Supporting Cell Testicular Dysgenesis Syndrome Testing Testis Testosterone Time Tissues Transgenes United States National Institutes of Health X Chromosome blastomere structure cell motility cell type cofactor experimental study fetal fetus cell gene discovery gene product gonad development idiopathic infertility in vivo interest knock-down knockout animal leydig interstitial cell male male fertility member mutant novel postnatal promoter protein function reproductive selective expression self-directed learning sertoli cell sex determination sperm cell stem stem cell niche subfertility tool transcription factor transcriptome transcriptome sequencing

项目摘要

PROJECT SUMMARY/ABSTRACT The Reproductive Homeobox X-linked (Rhox) genes encode a novel family of transcription factors that are excellent candidates to regulate events during spermatogenesis and ovulation. The RHOX homeodomain protein family has several members (13 distinct genes, with some existing in multiple copies). However, only two are expressed in Sertoli cells, the master control cells that promote testis development and nurse the differentiation and survival of germ cells. Ablation of these two genes, Rhox5 and Rhox8, results in male subfertility characterized by lower epididymal sperm numbers, motility defects, and reduced fecundity. Progeny from Rhox5-null mice, crossed with RHOX8 in vivo RNAi knockdown mice possess a distinct phenotype that is more severe than either single mutant. While these two mouse lines are useful tools for discovering the role of RHOX5 and RHOX8 after puberty, they are incapable of shedding light on RHOX8’s potential role in gonad development as the inhibitory Rhox8 transgene does not turn on until after post-natal day 7 when androgen signaling normally turns on Rhox5. To address this issue, a new complete Rhox8 deletion model will be employed to assess RHOX8’s function in embryonic Sertoli cell specification and function. Characterization of these mice is of great interest because: 1) Rhox8 is the only Rhox gene expressed in Sertoli cells of the embryonic gonad. Thus, there will be no concerns of functional redundancy which has been an issue for Rhox studies in the past. All of the other members of the cluster are restricted to primordial germ cells, including Rhox5 which is only found in Sertoli cells postnatally. 2) Analysis of the Rhox8 postnatal in vivo knockdown mic, and transient knockdown of RHOX8 in embryonic testis cultures, show that RHOX8 regulates Sox9. 3) SOX9 is a master regulator downstream of SRY – the factor that specifies male vs. female differentiation. Thus, RHOX8 may be in the sex determination pathway between SRY and SOX9. 4) After sex-determination, SOX9 governs multiple key events in the development of the tests including the formation of testis cords, male- specific vasculature, and differentiation of Sertoli and Leydig cell populations. If these post-puberty relationships are also conserved in the embryo, then RHOX8 may be a key cofactor or regulator of several events during gonad development. The experiments outlined in this proposal seek to systematically position Rhox8 in the milieu of male sexual development and discover the gene networks that are under its control. Because RHOX5 and RHOX8 may possess complementary functions in postnatal Sertoli cells, we will examine spermatogenesis in Rhox5/Rhox8 double knockout animals. Preliminary data suggests that these mice have a distinct and more severe phenotype than either single mutant. The elucidation of Rhox8’s function in Sertoli cells is important because it will provide an important “building block” towards the long-term goal of learning the independent and collaborative functions of all the Rhox genes in the testes.

项目概要/摘要生殖同源框 X 连锁 (Rhox) 基因编码一个新的转录因子家族，是调节精子发生和排卵过程中事件的优秀候选者。蛋白质家族有多个成员（13 个不同的基因，其中一些存在多个拷贝）。其中两种在支持细胞中表达，支持细胞是促进睾丸发育和滋养睾丸的主要控制细胞。 Rhox5 和 Rhox8 这两个基因的消融导致生殖细胞的分化和存活。生育力低下的特征是附睾精子数量减少、运动缺陷和生育力降低。来自 Rhox5 缺失小鼠，与 RHOX8 体内 RNAi 敲低小鼠杂交，具有独特的表型虽然这两个小鼠品系是发现的作用的有用工具，但比任何一个单一突变体都更严重。青春期后的 RHOX5 和 RHOX8 无法揭示 RHOX8 在性腺中的潜在作用由于抑制性 Rhox8 转基因直到产后第 7 天雄激素释放后才启动信号通常会开启 Rhox5。为了解决这个问题，我们将推出一个新的完整的 Rhox8 删除模型。用于评估 RHOX8 在胚胎支持细胞规格和功能表征中的功能。这些小鼠引起了极大的兴趣，因为：1）Rhox8 是唯一在支持细胞中表达的 Rhox 基因。因此，不会出现功能冗余的问题，而功能冗余一直是 Rhox 的问题。过去的研究仅限于该簇的所有其他成员，包括原始生殖细胞。 Rhox5 仅在出生后的支持细胞中发现。 2) 出生后体内敲低的 Rhox8 分析。 mic 和胚胎睾丸培养物中 RHOX8 的瞬时敲低表明 RHOX8 调节 Sox9 3)。 SOX9 是 SRY 下游的主要调节因子，SRY 是决定男性与女性差异的因素。因此，RHOX8 可能位于 SRY 和 SOX9 之间的性别决定途径中。 4) 性别决定后， SOX9 控制着测试开发中的多个关键事件，包括睾丸索的形成、男性- 特定的脉管系统，以及支持细胞和间质细胞群的分化，如果这些是青春期后的。关系在胚胎中也是保守的，那么 RHOX8 可能是几个关键的辅助因子或调节因子本提案中概述的实验旨在系统地定位性腺发育过程中的事件。 Rhox8 在男性性发育的环境中并发现受其控制的基因网络。由于 RHOX5 和 RHOX8 在出生后支持细胞中可能具有互补的功能，我们将检查 Rhox5/Rhox8 双敲除动物的精子发生，初步数据表明这些。与任一单一突变体相比，小鼠具有独特且更严重的表型。支持细胞的功能很重要，因为它将为长期发展提供重要的“基石” 目标是在测试中了解所有 Rhox 基因的独立和协作功能。