Neuronal control of body weight and size by the Gbeta5-R7 signaling complex

Gbeta5-R7 信号复合物对体重和体型的神经元控制

基本信息

批准号：
10001791
负责人：
Vladlen Z Slepak
金额：
$ 19.19万
依托单位：
UNIVERSITY OF MIAMI SCHOOL OF MEDICINE
依托单位国家：
美国
项目类别：
财政年份：
2019
资助国家：
美国
起止时间：
2019-09-15 至 2020-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10001791
关键词：
Ablation Acute Adult Affect Agonist Alleles Animals Antibodies Behavior Beta Cell Biochemical Biological Assay Biophysics Body Size Body Weight Brain CRISPR/Cas technology Cardiovascular Diseases Cardiovascular system Cell Line Cell Nucleus Cells Comorbidity Complex Control Animal Data Diabetes Mellitus Diagnostic Disease Energy Intake Engineering Etiology Event Family G Protein-Coupled Receptor Signaling G-Protein-Coupled Receptors GNB5 gene GTP-Binding Protein Regulators GTP-Binding Protein beta Subunits GTP-Binding Proteins Gene Transfer Genes Genetic Goals Growth Health Hormone secretion Hormones Human Human Genetics Hypothalamic Hormones Hypothalamic structure Image Analysis In Situ In Vitro Injections Insulin Investigation Kidney Diseases Knock-out Lead Link LoxP-flanked allele Malignant Neoplasms Measurement Mediating Messenger RNA Metabolic Diseases Metabolic syndrome Metabolism Methods Microscopic Molecular Mus Muscarinic Acetylcholine Receptor Mutation Neuraxis Neuroendocrinology Neuroglia Neurologic Neurons Neurosciences Neurosecretion Neurosecretory Systems Neurotransmitters Non-Insulin-Dependent Diabetes Mellitus Obesity Organ Pancreas Pathologic Pathway interactions Pattern Peripheral Pharmacology Phenotype Phosphorylation Physiological Play Population Proteins RGS Proteins Regulation Reproduction Research Risk Factors Role Second Messenger Systems Serum Signal Transduction Signaling Protein Slice Stimulus Stroke Structure Structure of beta Cell of islet System Technology Testing Therapeutic Tissues United States Viral Viral Vector Virus Weight Gain base cholinergic design dimer effective therapy experimental study hormonal signals hormone regulation improved in vivo insight insulin secretion kinase inhibitor knock-down knockout gene mutant nervous system disorder novel overexpression promoter protein complex protein function protein protein interaction real-time images

项目摘要

Abstract Obesity is a major co-morbidity factor in diabetes, cancer, cardiovascular, neurological and other diseases. One group of genes recently linked to obesity in humans are genes encoding the R7 family of regulator of G protein signaling proteins (RGS6, 7, 9 and 11) and Gnb5, which encodes the G protein beta subunit G5. G5 and R7 proteins form obligatory dimers, so that the Gnb5 knockout causes complete degradation of the entire G5-R7 complex. This proposal studies how a deficiency in the Gnb5 and R7 genes can lead to obesity; 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. 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 G5-R7 complex in pancreatic beta cells where G5-R7 strongly promoted insulin secretion. Brain is the main organ regulating body weight, and the expression level of G5-R7 is much higher than in the pancreas or any other peripheral tissues. These considerations lead to the hypothesis that G5-RGS7 regulates body weight via its function in the CNS, where as in the insulin-secreting beta cells, G5-R7 can regulate hormone and/or neurotransmitter secretion by neurons. In support of this hypothesis, recent studies in the lab demonstrated that local knockout of Gnb5 in the hypothalamus using LoxP/Cre technology caused a dramatic increase of body weight and adiposity in adult mice. The proposed research will develop this discovery. Specific Aim 1 will analyze the effects of Gnb5 ablation in specific hypothalamic nuclei and types of neurons on body weight and levels of various hormones. Overexpression of Gnb5 using viral-mediated gene transfer will be performed to rescue obesity. Aim 2 will determine which hormones and signaling proteins (i.e., GPCRs) are co-expressed with Gnb5 in hypothalamic neurons. Tissue and acutely cultured neurons will be examined for secretion of specific hormones and signaling. Specific Aim 3 will study molecular events affected by G5-R7 via its knockout (CRISPR/Cas9), knockdown and overexpression in cell lines that endogenously express M3R and G5-R7. Experiments will focus on signaling, structure-function analysis of G5-R7 and signal-stimulated secretion utilizing pharmacological agents and biochemical and biophysical assays. Through unraveling a novel mechanism that regulates neuronal functions, this project may have a strong impact on understanding etiology of obesity and other neuroendocrine disorders.

抽象的肥胖是糖尿病、癌症、心血管、神经系统和其他疾病的主要共病因素最近与人类肥胖相关的一组基因是编码 R7 家族的基因。 G 蛋白信号蛋白（RGS6、7、9 和 11）和 Gnb5（编码 G 蛋白 beta）的调节因子亚基 G5。G5 和 R7 蛋白形成必需二聚体，因此 Gnb5 敲除导致完全敲除。该提案研究了 Gnb5 和 R7 基因的缺陷如何影响整个 G5-R7 复合物的降解。可能导致肥胖；最终目标是找到与肥胖相关的疾病的有效治疗方法神经内分泌途径受这些基因调节。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强有力的前提在这项研究中，消除一个 Gnb5 等位基因会导致小鼠肥胖和代谢综合征；后来通过人类遗传学的证实，该实验室的后续机制研究发现了一种新的作用。胰腺β细胞中的G5-R7复合物，其中G5-R7强烈促进胰岛素分泌。是调节体重的主要器官，G5-R7的表达水平远高于胰腺或胰腺这些考虑导致了 G5-RGS7 调节身体的假设。通过其在 CNS 中的功能来调节体重，与分泌胰岛素的 β 细胞一样，G5-R7 可以调节激素和/或神经元分泌的神经递质支持这一假设，最近的实验室研究。证明使用 LoxP/Cre 技术局部敲除下丘脑中的 Gnb5 会导致显着的成年小鼠体重和肥胖的增加将进一步发展这一发现。具体目标 1 将分析 Gnb5 消融对特定下丘脑核团和神经元类型的影响使用病毒介导的基因转移来过度表达 Gnb5 会影响体重和各种激素的水平。目标 2 将确定哪些激素和信号蛋白（即 GPCR）是拯救肥胖的。将检查组织和急性培养的神经元中与 Gnb5 的共表达。具体目标 3 将研究受 G5-R7 影响的分子事件。通过在内源性表达 M3R 的细胞系中进行敲除 (CRISPR/Cas9)、敲低和过表达和 G5-R7 实验将重点关注 G5-R7 的信号传导、结构功能分析和信号刺激。利用药物制剂和生物化学和生物物理测定法进行分泌。调节神经功能的新机制，该项目可能会对理解产生重大影响肥胖和其他神经内分泌疾病的病因学。