Molecular Bases Committing Primate Spermatogonia to a Pathway of Differentiation.

描述（由申请人提供）：本项目的长期目标是了解控制恒河猴（一种代表性的高等灵长类动物）精原细胞分化的细胞和分子机制。该建议的基本策略是基于，在某种程度上，发现，而精原细胞增殖发生在促性腺激素刺激的情况下，灵长类动物精原细胞的分化只发生在促性腺激素驱动的存在下。这两个过程的内分泌需求的差异将被用来询问睾丸基因，基因网络和任何新的转录，决定精原细胞更新或分化的决定。有三个主要问题。首先，精原细胞内的哪些基因和途径决定了细胞的分化而不是自我更新。第二，什么是促性腺激素依赖性支持细胞旁分泌信号，指导未分化的精原细胞进行分化的路径。第三，精原细胞分化在多大程度上依赖于维生素A。为了开始回答这些问题，RNA-seq将用于评估生殖细胞和支持细胞基因表达和转录组谱的总体变化，这些变化与将未分化的A型精原细胞置于促性腺激素依赖性分化途径的初始步骤相关。在通过qRTPCR确认后，将使用免疫组织化学（IHC）和/或原位杂交（ISH）确定通过qRTPCR显示与精原细胞分化相关的上调或下调的那些基因和相关转录物的细胞特异性表达。将通过给动物喂食维生素A缺乏饲料来确定维生素A缺乏（VAD）对精原细胞增殖的影响。将讨论两个特定目的：1.鉴定与未分化的Ap型精原细胞决定进入分化途径相关的生殖细胞和支持细胞基因、基因网络和新转录物; 2.确定猴中未分化的Ap型精原细胞决定进入分化途径是否依赖于维生素A。这项工作将在一个互动的智力环境中进行，该环境由一组对精原生物学和支持细胞功能相关方面感兴趣的研究人员组成。预计拟议的工作将提供基本的见解精原细胞分化：精子发生的一个重要方面，将有关的治疗男性不育症的人类和新的男性避孕药的发展。

DESCRIPTION (provided by applicant): The long-term goal of this project is to understand the cellular and molecular mechanisms that control spermatogonial differentiation in the rhesus monkey, a representative higher primate. The strategy underlying the proposal is based, in part, on the finding that while spermatogonial proliferation occurs in the absence of gonadotropin stimulation, differentiation of primate spermatogonia occurs only in the presence of a gonadotropin drive. This difference in the endocrine requirements of the two processes will be used to interrogate the testicular genes, gene networks and any novel transcripts that determine the decision of spermatogonia to renew or to differentiate. There are three major questions. First, what genes and pathways within spermatogonia underlie the decision of the cell to differentiate rather than undergo self-renewal. Second, what are the gonadotropin dependent Sertoli cell paracrine signals that instruct undifferentiated spermatogonia to proceed down the path of differentiation. Third, to what extent is spermatogonial differentiation dependent on Vitamin A. To begin to answer these questions, RNA-seq will be used to assess global changes in germ cell and Sertoli cell gene expression and transcriptome profiles associated with the initial step in placing undifferentiated type A spermatogonia on the gonadotropin dependent pathway of differentiation. Following confirmation by qRTPCR, cell specific expression of those genes and relevant transcripts that are shown by qRTPCR to be up- or down-regulated in association with spermatogonial differentiation will then be determined using immunohistochemistry (IHC) and/or in situ hybridization (ISH). The effects of Vitamin A deficiency (VAD) on spermatogonial proliferation will be determined by feeding the animals a Vitamin A deficient diet. Two Specific Aims will be addressed; 1 to identify germ cell and Sertoli cell genes, gene networks and novel transcripts associated with the decision of undifferentiated type Ap spermatogonia to commit to a pathway of differentiation, and 2 To determine whether the decision of undifferentiated Type Ap spermatogonia to commit to the path of differentiation in the monkey is dependent on Vitamin A. This work will be conducted in an interactive intellectual environment that is generated by a group of investigators interested in related aspects of spermatogonial biology and Sertoli cell function. It is anticipated that the proposed work will provide fundamental insight into spermatogonial differentiation: a critical aspect of spermatogenesis that will be relevant to the treatment of infertility in the human male and to the development of novel male contraceptives.