The role of regulator of G protein signaling Gbeta5-R7 in neuronal control of body weight

肥胖是糖尿病、癌症、心血管疾病、神经系统疾病和其他疾病的主要共病因素。 疾病最近，一组新的人类基因与肥胖有关，这些基因编码R7家族 G蛋白信号传导蛋白（RGS 6、7、9和11）和Gnb 5的调节因子，后者编码非典型G 蛋白β亚基Gb 5。该提案研究Gnb 5和R7基因的缺陷如何导致 肥胖和其他体重控制异常。最终目标是发现有效的 治疗与这些基因调节的神经内分泌途径相关的疾病。 拟议的研究计划建立在早期发现的基础上，这些发现共同构成了一个强大的 本研究的前提。Gb 5和R7蛋白形成专性二聚体，因此Gnb 5敲除导致 整个Gb 5-R7复合物完全降解。一个Gnb 5等位基因的消除导致肥胖， 小鼠代谢综合征;这一发现后来被人类遗传学证实。后继机械论 本实验室的研究确定了Gb 5-R7复合物在胰腺β细胞中的新作用，其中Gb 5-R7复合物在胰腺β细胞中的作用是： R7强烈促进胰岛素分泌。脑是调节体重的主要器官， Gb 5-R7的表达水平在CNS中比在胰腺或任何其它外周组织中高得多。 这些考虑导致我们假设Gb 5-RGS 7通过其在以下方面的功能调节体重： 神经元，类似于分泌胰岛素的β细胞，它控制神经递质的分泌 和荷尔蒙。为了支持这一假设，最近的实验室研究表明， 成年小鼠下丘脑中的gnb 5显著增加了身体和肥胖。拟议 实验将通过在生物体上进行的一系列实验来发展这一发现， 组织和细胞水平。具体目标1将使用Cre-loxP方法在具体的 下丘脑核和神经元类型，并检查敲除对体重的影响， 新陈代谢.将使用病毒基因转移进行Gnb 5的过表达以挽救变化 是由击倒造成的目的2研究Gnb 5与GPCR的共表达及GPCR基因的编码 将离体检查下丘脑激素和组织切片的信号刺激分泌， 特定的荷尔蒙Specific Aim 3将研究Gb 5-R7及其敲除（CRISPR/Cas9）、敲减和 在内源性表达Gb 5-R7的细胞系中过量表达。实验将利用药理学 试剂和结构-功能分析，以询问信号传导和分泌途径中的分子事件。 对Gb 5-R7复合物的研究将阐明调节分泌的新机制 下丘脑激素和显着推进我们的理解G蛋白信号，病因， 肥胖和其他代谢和神经内分泌紊乱。

Obesity is a major co-morbidity factor in diabetes, cancer, cardiovascular, neurological and other diseases. Recently, a new group of human genes was linked to obesity, the genes encoding the R7 family of regulator of G protein signaling proteins (RGS6, 7, 9 and 11) and Gnb5, which encodes the atypical G protein beta subunit Gb5. This proposal studies how a deficiency in the Gnb5 and R7 genes can lead to obesity and other abnormalities in control of body weight. The ultimate goal is to discover effective treatments of disorders associated with neuroendocrine pathways regulated by these genes. The proposed research plan builds upon earlier discoveries that together constitute a strong premise for this study. Gb5 and R7 proteins form obligatory dimers, so that the Gnb5 knockout causes complete degradation of the entire Gb5-R7 complex. Ablation of one Gnb5 allele leads to obesity and metabolic syndrome in mice; this finding was later confirmed by human genetics. Subsequent mechanistic investigations in this lab identified a novel role for the Gb5-R7 complex in pancreatic beta cells where Gb5- R7 strongly promoted secretion of insulin. Brain is the main organ regulating body weight, and the expression level of Gb5-R7 is much higher in the CNS than in the pancreas or any other peripheral tissues. These considerations lead us to the hypothesis that Gb5-RGS7 regulates body weight via its function in the neurons, where, similarly to the insulin-secreting beta cells, it controls secretion of neurotransmitters and hormones. In support of this hypothesis, recent studies in the lab demonstrated that local knockout of Gnb5 in the adult mouse hypothalamus dramatically increased body and adiposity. The proposed experiments will develop this discovery through a series of experiments performed at the organismal, tissue and cellular levels. Specific Aim 1 will use the Cre-loxP approach to inactivate Gnb5 in specific hypothalamic nuclei and types of neurons and examine the effect of the knockout on body weight and metabolism. Overexpression of Gnb5 using viral gene transfer will be performed to rescue the changes caused by the knockout. Aim 2 will study co-expression of Gnb5 with GPCRs and gene encoding hypothalamic hormones and tissue slices will be examined ex vivo for signal-stimulated secretion of specific hormones. Specific Aim 3 will study Gb5-R7 its knockout (CRISPR/Cas9), knockdown and overexpression in cell lines that endogenously express Gb5-R7. Experiments will utilize pharmacological agents and structure-function analysis to interrogate molecular events in signaling and secretory pathways. The proposed investigation of the Gb5-R7 complex will elucidate the novel mechanism regulating secretion of hypothalamic hormones and significantly advance our understanding of G protein signaling, etiology of obesity and other metabolic and neuroendocrine disorders.