Transcription & Function of Adipocyte Beta3AR in Obesity

β 3-肾上腺素能受体（β 3 AR）是一种主要在脂肪组织中表达的G蛋白偶联受体（GPCR），在脂肪组织中介导去甲肾上腺素刺激的白色脂肪组织（WAT）脂解作用和棕色脂肪组织（BAT）产热作用。 β 3AR的选择性激动剂可以预防或逆转犬和啮齿动物模型中的肥胖和胰岛素抵抗，并且正在进行用于人类的药物开发。 除了有效刺激脂解外，β 3 AR激动剂还促进典型白色脂肪库中产热活性棕色脂肪细胞的出现，但这种作用的机制尚不清楚。 在我们之前的资助期内完成的目标确定，脂肪细胞中的β 3 AR与Galphas和Galphai组成性偶联，分别导致蛋白激酶A（PKA）和MAP激酶（ERK 1/2）通路的激活，并且β 3 AR的脂肪细胞特异性表达在很大程度上由C/EBPalpha控制，我们定义了C/EBP位点。 在探索β 3 AR对脂肪细胞中PKA和MAPK通路的调节中，我们最近发现β AR以PKA依赖的方式激活脂肪细胞中的p38 MAPK激酶，并且这种调节的重要生理终点之一是诱导棕色脂肪中的主要产热蛋白UCP 1。提出了两个主要假设。 从PKA到MKK 3/6和p38的信号级联涉及Rac 1/cdc 42和混合谱系激酶（MLK），并且p38 MAPK途径需要与PKA串联以适当地激活和募集转录因子到UCP 1启动子，最终终点是增加产热。 提出了三个目标：（1）表征脂肪细胞中β AR与p38 MAP激酶之间的信号转导途径;检验Rac 1/Cdc 42和MLK被PKA激活，进而直接激活p38 MAPK的上游激酶MKK 3/6的假设;（2）检验β 3AR刺激p38 MAP激酶活性调节脂肪细胞中PPAR依赖性转录过程的假设。 我们将测试PPARgamma和/或PGC-1的磷酸化是否是UCP 1基因反式激活的必需步骤，并测试需要PKA直接和间接调节CREB和PPARgamma的双重信号级联触发UCP 1转录的假设;（3）为了检验孤儿受体ROR γ是组织的第二关键转录激活因子的假设，β 3 AR基因在脂肪细胞中的特异性表达。 在此目的的实验也将测试的假设，在C/EBP蛋白质的轮廓的改变有助于肥胖症/糖尿病依赖性的β 3 AR和β 1 AR基因在脂肪细胞中的表达下调。

The beta3-adrenergic receptor (beta3AR) is a G protein- coupled receptor (GPCR) expressed predominantly in adipose tissue, where it mediates the effects of noradrenaline-stimulated lipolysis in white adipose tissue (WAT) and thermogenesis in brown adipose tissue (BAT). Selective agonists for the beta3AR can prevent or reverse obesity and insulin resistance in canine and rodent models, and are under pharmaceutical development for humans. In addition to potent stimulation of lipolysis, beta3AR agonists promote the appearance of thermogenically active brown adipocytes in typical white adipose depots but the mechanism responsible for this effect is not understood. Completion of the Aims in our previous funding period established that the beta3AR in adipocytes is constitutively coupled to both Galphas and Galphai, leading to the activation of the protein kinase A (PKA) and MAP kinase (ERK1/2) pathways, respectively, and that the adipocyte-specific expression of beta3AR is controlled in large part by C/EBPalpha and we defined the C/EBP site. In probing the regulation of the PKA and MAPK pathways in adipocytes by beta3AR, we have recently discovered that betaARs activate p38 MAPK kinase in adipocytes in a PKA-dependent manner, and that one of the important physiological endpoints of this regulation is the induction of the main thermogenic protein in brown fat, UCP1. Two main hypotheses are proposed. That the signaling cascade from PKA to MKK3/6 and p38 involves Rac1/cdc42 and the mixed- lineage kinases (MLKs), and that the p38 MAPK pathway is required in tandem with PKA to appropriately activate and recruit transcription factors to the UCP1-promoter, with the ultimate end point being increased thermogenesis. There are three Aims proposed: (1) to characterize the signal transduction pathway linking the betaARs in adipocytes to the p38 MAP kinase; to test the hypothesis that Rac1/Cdc42 and the MLKs are activated by PKA and, in turn, directly activate MKK3/6, the immediate upstream kinase for p38 MAPK; (2) to test the hypothesis that beta3AR- stimulation of p38 MAP kinase activity regulates PPAR-dependent transcriptional processes in adipocytes. We will test whether phosphorylation of PPARgamma and/or PGC-1 are required steps in the transactivation of the UCP1 gene, and test the hypothesis that a dual signaling cascade requiring PKA both directly and indirectly to modulate CREB and PPARgamma triggers UCP1 transcription; (3) to test the hypothesis that the orphan receptor RORgamma is a second critical transcriptional activator for the tissue-specific expression of the beta3AR gene in adipocytes. Experiments in this aim will also test the hypothesis that alterations in the profile of C/EBP proteins contribute to the obesity/diabetes dependent down-regulation of expression of the beta3AR and beta1AR genes in adipocytes.