Transcriptional role of TLE3 in brown adipose tissue development and metabolism

描述（由申请人提供）：肥胖会增加患慢性疾病的风险，如2型糖尿病、心血管疾病和癌症。肥胖源于能量摄入和能量消耗之间的不平衡，其中多余的能量以甘油三酯的形式储存在脂肪细胞中。PPAR吗?是脂肪形成的关键调节剂，调节脂肪细胞谱系特征基因的表达。促进PPAR的机制是什么？脂肪形成过程中依赖基因的表达尚不完全清楚。之前我们报道过TLE3是PPAR？它参与了一个前馈转录程序来驱动脂肪形成。在拟议的研究中，我将验证TLE3是驱动白色脂肪与棕色脂肪选择性基因表达的关键决定因素的假设。初步数据表明，在棕色脂肪组织（BAT）中过表达TLE3的小鼠具有从棕色脂肪组织到白色脂肪组织（WAT）的表型转换。结果，TLE3转基因小鼠在低温暴露时产热反应受损。机制研究表明，TLE3对抗Prdm16，一种棕色脂肪基因表达的转录共调节因子。在具体的Aim 1中，我将使用体外模型来研究TLE3在执行WAT和BAT转录程序中的功能和作用机制。我将利用体外功能增益和损失研究来确定TLE3是否影响白色脂肪和棕色脂肪基因表达。功能途径的增加将包括逆转录病毒和腺病毒表达TLE3和/或Prdm16。功能丧失研究将利用感染对照或cree腺病毒的白色和棕色TLE3F/F前脂肪细胞产生体外敲除。我将通过检测TLE3引导染色质重塑和组蛋白修饰酶募集到脂肪细胞启动子的能力来探索其作用机制。在具体的Aim 2中，我将使用体内模型来定义TLE3在白色和棕色脂肪组织和全身脂质代谢中的功能。我已经培育出在脂肪细胞中表达TLE3的转基因动物，以及在脂肪细胞中条件缺失TLE3的小鼠。我将使用这些模型来检验TLE3影响脂肪细胞基因表达、产热以及脂质和葡萄糖代谢的能力。拟议的研究将在加州大学洛杉矶分校Peter Tontonoz博士的实验室完成，他提供了一个丰富的环境，将有助于过渡到独立研究者的位置。职业发展奖将为培养写作、网络和沟通方面的关键技能提供保护时间。除了…之外

DESCRIPTION (provided by applicant): Obesity increases the risk for chronic diseases such as type 2 diabetes, cardiovascular disease, and cancer. Obesity results from the imbalance between energy intake and energy expenditure, where excess energy is stored in adipocytes as triglycerides. PPAR? is a critical regulator of adipogenesis, regulating the expression of genes that are characteristic of the adipocyte lineage. The mechanisms that facilitate PPAR?- dependent gene expression during the course of adipogenesis are incompletely understood. Previously we reported that TLE3 is a transcriptional coregulator of PPAR? and is involved in a feed-forward transcriptional program to drive adipogenesis. In the proposed studies I will test the hypothesis that TLE3 is a key determinant in driving white versus brown fat selective gene expression. Preliminary data indicates that mice overexpressing TLE3 in brown adipose tissue (BAT) have a phenotypic switch from brown to white adipose tissue (WAT). As a result, TLE3 transgenic mice have an impaired thermogenic response when challenged with cold exposure. Mechanistic studies suggest that TLE3 counters Prdm16, a transcriptional coregulator of brown fat gene expression. In specific Aim 1 I will use in vitro models to investigate the function and mechanism of action of TLE3 in executing the WAT and BAT transcriptional programs. I will utilize in vitro gain and loss of function studies to determine whether TLE3 affects white versus brown fat gene expression. Gain of function approaches will include retroviral and adenoviral expression of TLE3 and/or Prdm16. Loss of function studies will utilize white and brown TLE3F/F preadipocytes infected with control or Cre-adenovirus to generate in vitro knockouts. I will explore the mechanism of action by examining the ability of TLE3 to direct chromatin remodeling and recruitment of histone modifying enzymes to adipocyte promoters. In specific Aim 2 I will use in vivo models to define the function of TLE3 in white and brown adipose tissue and systemic lipid metabolism. I have already generated transgenic animals expressing TLE3 in adipocytes, as well as mice with conditional deletion of TLE3 in adipocytes. I will use these models to examine the ability of TLE3 to affect adipocyte gene expression, thermogenesis and lipid and glucose metabolism. The proposed studies will be completed in the laboratory of Dr. Peter Tontonoz at UCLA, who has provided an enriching environment that will facilitate the transition to an independent investigator position. The career development award will provide protected time to develop critical skills in writing, networking, and communication. In addition to attending seminars at UCLA and attending conferences, I will take courses that will enhance technical training in the use of radioisotopes for the study of metabolic pathways, protein purification and characterization, and next generation sequencing technologies for the study of epigenetics. The proposed studies are a logical transition from my postdoctoral studies in adipogenesis to the burgeoning field of brown adipocyte biology.