Whole-genome analysis of direct Doublesex target genes

描述（由申请人提供）：创造不同的雄性和雌性形态（两性异形）是有性繁殖动物发展的关键一步。尽管启动性别决定的机制在不同物种之间差异很大，但最近的研究表明，性别决定控制产生两性二态现象的途径更为保守。一个很好的例子是双性Mab3相关转录因子（DMRT）家族，该家族已被证明在果蝇、蠕虫、鱼类、鸟类、小鼠和人类等多种生物中调节两性二态性。然而，这些关键转录因子控制两性二态性的分子途径在很大程度上仍未被探索。DMRT家族的创始成员是双性果蝇蛋白（DSX），它是控制果蝇雌雄形态的主要因素。dsx RNA在雄性和雌性中拼接成不同的形式，编码雄性和雌性蛋白DSXM和DSXF。这两种高度相关的转录因子如何调节相反的雄性和雌性发育程序是该领域的一个关键问题，但很少有DSX的靶点被确定。我们建议通过采用全基因组分子方法来揭示由DSXM和DSXF控制的调控网络来解决这个问题。我们将使用染色质免疫沉淀和高通量测序（ChIP-seq）来鉴定基因组中的DSX结合位点。然后，我们将这些数据与男性和女性的RNA表达谱进行比较，以确定DSX靶基因。这些研究将使我们能够确定DSX控制性腺两性二态性的调节网络，并解决有关DSX的雄性和雌性形式如何差异调节该网络的基本问题。这也将为未来研究DSXM和DSXF调控其靶点的机制以及这些靶点在控制性二态发育中所起的作用奠定坚实的基础。这项工作将帮助我们理解人类性发育障碍，例如那些影响dmrt的疾病，并将为理解转录因子的选择性剪接通常如何导致多种下游反应提供一个框架。

DESCRIPTION (provided by applicant): The creation of distinct male and female forms (sexual dimorphism) is a critical step in the development of sexually reproducing animals. Although the mechanisms that initiate sex determination vary considerably between different species, recent work demonstrates that the pathways that sex determination controls to create sexual dimorphism are more well-conserved. An excellent example is the Doublesex Mab3 Related Transcription Factor (DMRT) family, which have been shown to regulate sexual dimorphism in organisms as diverse as flies, worms, fish, birds, mice and man. However, the molecular pathways by which these key transcription factors control sexual dimorphism remain largely unexplored. The founding member of the DMRT family is the Drosophila protein Doublesex (DSX), which is the main factor controlling male vs. female morphology in flies. The dsx RNA is spliced into distinct forms in males vs. females, which encode the male and female proteins, DSXM and DSXF. How these two, highly-related transcription factors regulate the opposing male vs. female developmental programs is a key question in the field, but few targets for DSX have been identified. We propose to address this question by taking a whole-genome molecular approach to uncover the regulatory network(s) controlled by DSXM and DSXF. We will use chromatin immunoprecipitation followed by high throughput sequencing (ChIP-seq) to identify DSX binding sites in the genome. We will then compare these data to RNA expression profiles on males vs. females to identify DSX target genes. These studies will allow us to identify the regulatory network through which DSX controls gonad sexual dimorphism and address fundamental questions about how the male and female forms of DSX differentially regulate this network. This will also create a strong foundation for future work that focuses on the mechanisms that DSXM and DSXF use to regulate their targets, and what role these targets play in controlling sexually dimorphic development. This work will help us to understand human disorders of sexual development, such as those affecting DMRTs, and will provide a framework for understanding how the alternative splicing of transcription factors in general can lead to a diverse repertoire of downstream responses. PUBLIC HEALTH RELEVANCE: This work studies how the differences between the sexes are regulated during development. This is relevant to our understanding of human Disorders of Sex Development (DSDs), which cause defects in sexual differentiation and infertility.