Regulation of a Spermatogenesis-Specific Gene

描述（由申请人提供）：精子发生是一个动态的细胞分化过程，其特征是基因表达模式的剧烈变化。多年来，我们研究了精子发生特异性Pgk2基因作为一个严格调控的组织特异性基因的模型，该基因仅在减数分裂和减数分裂后的生精细胞中表达。我们利用独特的可获得的生精细胞谱系来获得在减数分裂前、减数分裂和减数分裂后阶段的特定生精细胞类型的纯化群体。这使我们能够检查与转录调控机制相关的分子参数，并确定导致Pgk2基因转录起始的分子事件的发育顺序。这使我们能够对这一监管过程中涉及的机制提出可检验的假设。在这个应用中，我们提出了实验来确定参与该基因调控的关键因素，表征该基因转录激活之前染色质组成和结构的变化，最后，评估该基因中组织特异性和基因特异性去甲基化结构域先于任何其他转录参数变化而形成的机制，并评估体内Pgk2基因的转录激活需要去甲基化的程度。这些实验将使我们能够定义与精子发生特异性基因转录激活相关的调节事件之间的因果关系。这将有助于我们更好地理解组织特异性基因表达的调控机制，尤其是精子发生特异性基因表达的调控机制。

DESCRIPTION (provided by applicant): Spermatogenesis is a dynamic process of cellular differentiation that is marked by dramatic changes in patterns of gene expression. For many years we have studied the spermatogenesis-specific Pgk2 gene as a model of a tightly regulated tissue-specific gene expressed exclusively in meiotic and postmeiotic spermatogenic cells. We have exploited the uniquely accessible spermatogenic cell lineage to obtain purified populations of specific spermatogenic cell types at premeiotic, meiotic, and postmeiotic stages. This has allowed us to examine molecular parameters associated with mechanisms of transcriptional regulation, and to determine a developmental order of molecular events that lead up to initiation of transcription of the Pgk2 gene. This has allowed us to formulate testable hypotheses about the mechanisms involved in this regulatory process. In this application, we propose experiments to identify key factors involved in the regulation of this gene, to characterize changes in chromatin composition and structure that precede transcriptional activation of this gene, and, finally, to assess the mechanism by which a tissue- and gene-specific demethylation domain develops in this gene prior to any other changes in transcriptional parameters and to assess the extent to which demethylation is required to predispose transcriptional activation of the Pgk2 gene in vivo. These experiments will allow us to define cause-and-effect relationships among regulatory events associated with transcriptional activation of a spermatogenesis-specific gene. This will lead to an enhanced understanding of the mechanisms that regulate tissue-specific gene expression in general, and spermatogenesis-specific gene expression in particular.