Regulation of Reproduction

该项目的长期目标是全面了解类固醇激素的合成是如何以发育和组织特异性的方式调节的，以及调节失调如何导致生殖疾病，如多囊卵巢综合征（PCOS）。甾体生成酶的基因在几个水平上受到转录调节：发育、组织特异性和激素。这种调节具有共同的特征，但在不同的类固醇酶基因和不同的哺乳动物之间也有所不同。我们将重点关注编码P450c17 （17α羟化酶/17,2裂解酶）的基因，因为它是甾体生成的关键分支点，它与PCOS的病因有关，并且可能参与神经和生殖系统的早期发育。它在人肾上腺中的表达是合成17羟基C21类固醇（如皮质醇）和性腺和大脑中合成C19性类固醇所必需的。在神经系统中，P450c17的表达也导致脱氢表雄酮的合成，这是一种有效的神经调节剂。我们一直使用啮齿动物作为我们的模型系统来研究该基因的转录调控，并已经确定了顺式作用的DNA元件和几个新的反式作用的核因子。其中一种转录因子，SET，已经从急性未分化白血病患者的染色体易位中鉴定出来，但其作为转录因子的作用尚不清楚。我们现在将通过鉴定其DNA结合、反活化和潜在的二聚化结构域，以及鉴定可能与SET相互作用参与其反活化功能的辅助因子来确定SET的作用机制。由于SET在发育性腺和神经系统中大量表达并调节P450c17的表达，我们将确定这些辅助因子是否在组织中特异性表达，和/或它们是否修饰SET的其他已知功能。我们在大鼠P450c17基因中定位顺式活性元件的研究已经确定了一个被我们称为StF-IT-2的因子结合的区域，该因子以一种新的方式与孤儿核受体SF-1相互作用，以调节P450c17的转录。我们将对这种相互作用进行表征，并纯化、表征和克隆StF-IT-2的cDNA。这些研究的成功完成将使我们更好地了解性腺和神经系统如何启动甾体生成，一类新的转录因子的作用机制，SF-1调节基因表达的新机制，并将确定另一个可能参与P450c17表达的独特方面的转录因子。

The long term objective of this project is a thorough understanding of how steroid hormone synthesis is regulated in a developmental and tissue-specific fashion, and how dysregulation may result in reproductive disorders such as polycystic ovarian syndrome (PCOS). The genes for steroidogenic enzymes are transcriptionally regulated at several levels: developmentally, tissue specifically, and hormonally. This regulation shares common features but also differs among the various genes for the steroidogenic enzymes and among various mammals. We will focus on the gene encoding P450c17 (17alpha hydroxylase/17,2 lyase) as it is the key branch point in steroidogenesis, it has been implicated in the etiology of PCOS, and it may be involved in early development of nervous and reproductive systems. Its expression in the human adrenal is required for the synthesis of 17 hydroxy C21 steroids (e.g. cortisol) and for synthesis of C19 sex steroids in the gonads and brain. In the nervous system, expression of P450c17 also results in DHEA synthesis, a potent neuromodulators. We have been using the rodent as our model system for studying the transcriptional regulation of this gene, and have identified cis-acting DNA elements and several novel trans-acting nuclear factors. One of these transcription factors, SET, had been identified from a chromosomal translocation in a patient with acute undifferentiated leukemia, but its role as a transcription factor way unknown. We shall now determine the mechanism of SET action by identifying its DNA binding, transactivation, and potential dimerization domains, and identifying co-factors that may interact with SET to participate in its transactivating functions. AS SET is abundantly expressed and regulates P450c17 expression in the developing gonad and nervous system, we will determine if these co-factors are tissue- specifically expressed, and/or if they modify other known functions of SET. Our studies localizing cis-active elements in the rat P450c17 gene have identified a region bound by a factor that we call StF-IT-2, that interacts with the orphan nuclear receptor SF-1 in a novel way, to regulate P450c17 transcription. We will characterize that interaction, and purify, characterize and clone the cDNA for StF-IT-2. Successful completion of these studies will give us a better understanding of how the gonadal and nervous systems initiate steroidogenesis, the mechanism of action of a new class of transcription factor, novel mechanisms by which SF-1 regulates gene expression, and will identify another transcription factor that may participate in a unique aspect of P450c17 expression.