Regulation of Growth Factors and Embryogenesis

描述（由申请人提供）：本研究的目的是扩大我们对胚胎发生过程中起关键作用的基因调控网络的理解。最近的研究表明，转录因子Sox2和Oct-3/4都表现出调控哺乳动物胚胎发生和胚胎干细胞自我更新的重要特征。Sox2和Oct-3/4协同工作，协调至少6个基因（FGF-4、UTF1、Fbx15、Nanog、Sox2和Oct-3/4）在胚胎发育期间和胚胎干细胞中表达的转录。这些基因被称为Sox2:Oct-3/4靶基因。这里提出的工作是基于3个新的发现和假设。首先，我们最近确定了19个可能是Sox2:Oct-3/4基因调控网络一部分的额外基因。鉴于已知的6个Sox2:Oct-3/4靶基因中有4个（可能有5个）对胚胎发生至关重要，我们的中心假设是，大多数Sox2:Oct-3/4靶基因在胚胎发生过程中起着关键作用，其中许多是胚胎干细胞自我更新所必需的。其次，已知的6个Sox2:Oct-3/4靶基因的表达被证明非常严重地依赖于每个基因增强子中Sox2和Oct-3/4紧密间隔的DNA结合位点。这些和其他的研究使我们假设Sox2:Oct-3/4基因调控网络是由一组共享的共激活子协调控制的，至少其中一些在分化过程中自己受到调节，因此是决定细胞命运的关键参与者。第三，虽然已知Sox2与Oct-3/4结合可以刺激干细胞中Sox2:Oct-3/4靶基因的表达，但我们确定当Sox2过表达时，6个Sox2:Oct-3/4靶基因中至少有5个（包括Sox2基因本身）受到抑制。这些发现导致了我们的假设，即Sox2:Oct-3/4靶基因受到负反馈回路和正反馈回路的仔细调节，以确保这些基因在胚胎发生所需的水平上表达。本文提出了三个具体目标来检验这些假设。1)研究一组新发现的Sox2:Oct-3/4靶基因对胚胎干细胞自我更新和哺乳动物发育的影响。2)研究特异性共激活因子在Sox2:Oct-3/4靶基因表达中的作用。3)确定Sox2过表达抑制Sox2:Oct-3/4靶基因表达的分子机制。总之，这些研究将为协调胚胎发生和控制胚胎干细胞自我更新的关键基因调控网络提供机制理解。因此，这些研究将提供新的见解，并对发育生物学、再生医学和癌症生物学产生重大影响。

DESCRIPTION (provided by applicant): The goal of this study is to broaden our understanding of a gene regulatory network that plays key roles during embryogenesis. Recent studies argue strongly that the transcription factors Sox2 and Oct-3/4 each exhibit important hallmarks of master regulators that orchestrate mammalian embryogenesis and the self-renewal of embryonic stem (ES) cells. Sox2 and Oct-3/4 have been shown to work together cooperatively to coordinate the transcription of at least 6 genes (FGF-4, UTF1, Fbx15, Nanog, Sox2 and Oct-3/4) expressed during embryogenesis and by ES cells. These genes are referred to as Sox2:Oct-3/4 target genes. The work proposed here is based on 3 novel findings and hypotheses. First, we recently identified 19 additional genes that are likely to be part of the Sox2:Oct-3/4 gene regulatory network. Given that 4, and possibly 5, of the 6 known Sox2:Oct-3/4 target genes are essential for embryogenesis it is our central hypothesis that the majority of Sox2:Oct-3/4 target genes play critical roles during embryogenesis, and that many are required for the self-renewal of ES cells. Second, the expression of the 6 known Sox2:Oct-3/4 target genes was shown to be very heavily dependent on closely spaced DNA binding sites for Sox2 and Oct-3/4 in the enhancer of each gene. These and other studies led us to hypothesize that the Sox2:Oct-3/4 gene regulatory network is controlled coordinately by a shared set of co-activators, at least some of which are themselves regulated during differentiation and, thus, are key players in cell fate determination. Third, although Sox2, in combination with Oct-3/4, is known to stimulate the expression of Sox2:Oct-3/4 target genes in stem cells, we determined that at least 5 of the 6 Sox2:Oct-3/4 target genes, including the Sox2 gene itself, are inhibited when Sox2 is overexpressed. These findings led to our hypothesis that Sox2:Oct-3/4 target genes are regulated carefully by a negative, as well as a positive, feedback loop to ensure that these genes are expressed at levels required for embryogenesis. Three Specific Aims are proposed to test these hypotheses. 1) Examine a newly identified set of putative Sox2:Oct-3/4 target genes for their effects on the self-renewal of ES cells and mammalian development. 2) Examine the roles of specific co-activators in the expression of Sox2:Oct-3/4 target genes. 3) Determine the molecular mechanisms by which Sox2 overexpression inhibits the expression of Sox2:Oct-3/4 target genes. Together, these studies will provide a mechanistic understanding of a critical gene regulatory network that orchestrates embryogenesis and controls the self-renewal of ES cells. As such, these studies will provide novel insights and have significant impact on developmental biology, regenerative medicine and cancer biology.