Unique Roles of p160 Coactivators during Human Adipogenesis

描述(由申请人提供)： 在全世界几乎所有的社会中，肥胖症的发病率都在急剧增加。此外，内脏脂肪在腹部积聚，主要是白色脂肪组织，与发生2型糖尿病、心血管疾病和发病率增加的风险增加是一致的。肥胖的主要细胞基础是脂肪细胞(脂肪细胞)的大小和数量增加，并以细胞内脂滴中甘油三酯的增加为标志。脂肪细胞的成熟在很大程度上是由核受体(NR)的激素诱导所驱动的；然而，关于补充核受体活性的特定的早期途径以及随后在人类细胞中的脂质积累，人们知之甚少。最近的研究表明，pi 60NR辅调节因子(SRC-1、SRC-2、SRC-3)是协调相互作用的转录因子电路以驱动脂肪生成所必需的。这些辅调节因子(COR)的遗传消融会导致肥胖(SRC-1)和白色脂肪沉积减少(SRC-3，SRC-2)。此外，SRC-1/SRC-3双基因敲除小鼠表现出瘦削的表型。在结构上，pI60S在负责NR和转录因子相互作用的六个结构域中具有序列同源性。生化研究已将pI60s与过氧化物酶体增殖物激活受体-γ(PPAR)联系在一起，PPAR是脂肪形成的主要调节因子。这些线索表明，pI60s和一个时间元素之间可能存在功能重叠，该时间元素驱动它们与PPAR的串扰，从而促进脂肪生成。因此，我假设pi-60s在人类成脂过程中有多余的作用。在这项研究中，我提出了以下目的，以定量指标pi-60s在人类脂肪形成早期阶段的作用和功能重叠。目的1建立SRC-1、SRC-2和SRC-3在脂肪形成早期阶段的时空定位和表达水平。目标2将确定SRC-1的功能冗余， SRC-2和SRC-3在人类脂肪形成过程中的作用。我们将采用siRNA和高通量显微镜相结合的方法来定量评价分化过程中这些pi 60辅助调节因子之间的成脂相加、协同或拮抗作用。这项研究的目的是利用高通量显微镜(HTM)和图像分析，在单细胞框架内更全面地了解pi-60s在人类脂肪形成过程中的作用和协调。 与公共健康相关：这种方法允许快速、平行地询问激素的作用，从而更全面地了解调节人类脂肪细胞分化的机制。这一发现将为预防肥胖的发生和相关的临床结果提供新的干预策略。

DESCRIPTION (provided by applicant): The incidence of obesity is increasing dramatically in virtually all societies throughout the world. Further, accumulation of visceral fat, predominantly as white adipose tissue, in the abdominal regions is coincident with an elevated risk of developing type 2 diabetes, cardiovascular disease, and increased morbidity. The dominant cellular basis of obesity is increased fat cell (adipocyte) size and number and marked by accretion of triglycerides within intracellular lipid droplets. Adipocyte maturation is largely driven by the hormonal induction of nuclear receptors (NR); however, little is known about the specific, early pathways that complement NR activity and subsequent lipid accumulation in human cells. Recent studies have shown that the pi 60 NR coregulators (SRC-1, SRC-2, SRC-3) are requisite to coordinate a circuit of interacting transcription factors to drive adipogenesis. Genetic ablation of these coregulators (CoR) leads to obesity (SRC-1) and a reduction in white adipose deposition (SRC-3, SRC-2). Additionally, SRC-1/SRC-3 double knockout mice exhibit a lean phenotype. Structurally, the pi 60s share sequence homology within the six domains responsible for NR and transcription factor interactions. Biochemical studies have linked the pi 60s to peroxisome proliferator-activated receptor-gamma (PPAR), the master regulator of adipogenesis. These clues suggest the potential for a functional overlap between the pi 60s and a temporal element that drives their crosstalk with PPAR to promote adipogenesis. Therefore, I hypothesize that the pi 60s have redundant roles during the human adipogenic program. In this study, I propose the following aims to quantitatively index the role and functional overlap of the pi 60s during the early phases of human adipogenesis. Aim 1 will establish the spatiotemporal localization and expression levels occurring throughout the early phases of adipogenesis for SRC-1, SRC-2, and SRC-3. Aim 2 will determine the functional redundancy of SRC-1, SRC-2, and SRC-3 during human adipogenesis. A combinatorial siRNA and high throughput microscopy method will be applied to quantitatively index the adipogenic additivity, synergy, or antagonism between these pi 60 coregulators during differentiation. The objective of this fellowship is to obtain a more complete understanding of the roles and coordination of the pi 60s during human adipogenesis in a single cell framework using high throughput microscopy (HTM) and image analysis. PUBLIC HEALTH RELEVANCE: This approach allows for a rapid, parallel interrogation of hormone action leading to a more complete understanding of the mechanisms regulating human adipocyte differentiation. The findings will provide new intervention strategies to prevent the onset of obesity and the associated clinical outcomes.