Novel regulatory mechanisms in adipogenesis: role of the ES-cell transcription f

描述(申请人提供)：导致脂肪细胞表型的分化程序由一系列转录因子、CCAAT/增强子结合蛋白(C/EBPs)和核激素受体--过氧化体增殖物激活受体？？控制。(PPAR？)C/EBP？和C/EBPA在脂肪形成的早期迅速诱导，并随后驱动C/EBP？PPAR呢？表情。C/EBP？PPAR呢？可以在正反馈循环中相互诱导，并直接激活许多终末脂肪细胞分化的基因。PPARG被认为是脂肪形成的主要调节因子，因为它本身就可以诱导C/EBP？缺陷小鼠胚胎成纤维细胞(MEF)，而C/EBP？在没有PPAR的情况下无法推动分化。叠加在C/EBPs和PPAR？的这个框架上，我们最近发现了胚胎干细胞转录因子Zfp206及其剪接变异体ZScan10/201的瞬时表达。Zfp206的表达在融合时发生，随着表达切换到剪接变异体，表达在第3天减弱。剪接变异体缺乏完整的锌指DNA结合域，我们提出了作为显性负值的功能。我们的假设是，Zfp206的表达是维持分化潜能所必需的，随着PPARg的表达建立起终末分化，Zfp206的表达减弱。剪接变体ZScan的瞬时表达确保Zfp206效应被否定。为了验证我们的假设，我们将通过敲除和过度表达研究以及鉴定这种胚胎干细胞转录因子激活的基因来确定Zfp206在分化程序中的作用。此外，我们还将通过分析Zfp206的5‘侧翼区和第一内含子来确定Zfp206表达的调控机制。这些研究将提供关于成脂分化计划中一种新的调控系统的信息，此前人们认为该计划局限于胚胎干细胞。 与公共健康相关：脂肪组织在糖尿病和肥胖症的病理过程中发挥着关键作用，并已被公认为是一种动态的内分泌样组织，合成和分泌负责调节能量储存和能量消耗之间平衡的蛋白质。如果我们要识别新的诊断生物标记物和发现新的治疗方案，我们需要了解分化过程的调节以及这种复杂组织中分化表型的维持。已经证明，分化过程的控制主要取决于两个转录因子家族，C/EBPs和PPAR？我们最近发现了胚胎干细胞转录因子Zfp206及其剪接变异体ZScan10/201的瞬时表达，我们认为这将为分化过程带来额外的调控水平。我们的假设是，Zfp206的表达对于维持分化潜能是必不可少的，随着PPAR？表达的终末分化的建立，Zfp206的表达减弱。因此，研究Zfp206和Zscan10/201作用机制的拟议研究有望提供有关控制脂肪细胞分化的新靶点的信息，从而对肥胖和糖尿病有基本的见解。

DESCRIPTION (provided by applicant): The differentiation program leading to the adipocyte phenotype is controlled by a cascade of transcription factors, the CCAAT/enhancer binding proteins (C/EBPs) and a nuclear hormone receptor, peroxisome proliferator activated receptor ??? (PPAR?). C/EBP? and C/EBPa are rapidly induced in the early stage of adipogenesis and subsequently drive C/EBP? and PPAR? expression. C/EBP? and PPAR? can induce each other in a positive feedback loop and directly activate many of the genes for terminal adipocyte differentiation. PPARg is considered the master regulator of adipogenesis as it alone can induce differentiation in C/EBP? deficient murine embryonic fibroblasts (MEFs), while C/EBP? is incapable of driving differentiation in the absence of PPAR. Superimposed on this framework of C/EBPs and PPAR?, we have recently identified the transient expression of the embryonic stem cell transcription factor Zfp206 and its splice variant ZScan10/201. Zfp206 expression occurs at confluency and is attenuated by day 3 as expression switches to the splice variant. The splice variant lacks the entire zinc-finger DNA binding domain and we propose functions as a dominant negative. Our hypothesis is that expression of Zfp206 is essential for maintenance of the differentiation potential and as terminal differentiation is established with the expression of PPARg, Zfp206 expression is diminished. The transient expression of the splice variant ZScan insures Zfp206 effects are negated. To test our hypothesis we will define the role of Zfp206 in the differentiation program through the use of knock out and over expression studies as well as the identification of genes activated by this embryonic stem cell transcription factor. Additionally we will determine the mechanism by which Zfp206 expression is controlled through analysis of the 5'flanking region and first intron. The studies will yield information on a novel regulatory system in the adipogenic differentiation program previously thought to be localized to embryonic stem cells. PUBLIC HEALTH RELEVANCE: Adipose tissue plays a critical role with respect to the pathology of both diabetes and obesity and has become recognized as a dynamic endocrine-like tissue, synthesizing and secreting proteins responsible for regulation of the balance between energy storage and energy expenditure. If we are to identify new diagnostic biomarkers and discover new treatment options, we need to understand the regulation of the differentiation process as well as the maintenance of the differentiated phenotype in this complex tissue. Control of the differentiation process has been demonstrated to rest largely with two families of transcription factors, the C/EBPs and PPAR?. We have recently identified the transient expression of the embryonic stem cell transcription factor Zfp206 and its splice variant ZScan10/201, that we propose bring an additional level of regulation to the differentiation process. Our hypothesis is that expression of Zfp206 is essential for maintenance of the differentiation potential and as terminal differentiation is established with the expression of PPAR?, Zfp206 expression is diminished. Thus, the proposed studies examining the mechanism of action of Zfp206 and Zscan 10/201 are expected to yield information on a novel target for control of adipocyte differentiation and thus fundamental insights into obesity and diabetes.