Gbeta5-R7 Complex in Signaling and Hormone Secretion

Gbeta5-R7 复合物在信号传导和激素分泌中的作用

• 批准号: 8864017
• 负责人: Vladlen Z Slepak
• 金额: $ 42.21万
• 依托单位: UNIVERSITY OF MIAMI SCHOOL OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-04-01 至 2019-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8864017
• 关键词: Agonist; Animals; Area; Arrestins; Beta Cell; Biochemical Pathway; Biological Assay; Body Weight; Cell Line; Cell membrane; Cells; Complex; Data; Development; Diabetes Mellitus; Diagnostic; Disease; Drug Regulations; Drug effect disorder; Employee Strikes; Endocrine Glands; Etiology; Family; Foundations; Future; G Protein-Coupled Receptor Signaling; GTP-Binding Protein Regulators; GTP-Binding Proteins; Gene Expression; Gene Expression Profile; Gene Expression Regulation; Gene Proteins; Glucose; Growth; Guanosine Triphosphate Phosphohydrolases; Hormones; In Situ Hybridization; Insulin; Investigation; Ion Channel; Islets of Langerhans; Knock-out; Knockout Mice; Laboratories; Locomotion; Measures; Mediating; Metabolic Diseases; Metabolism; Modeling; Molecular; Mus; Muscarinic M3 Receptor; Muscarinics; Nerve; Nervous system structure; Neurons; Nutrient; Obesity; Pancreas; Pathway interactions; Phosphotransferases; Physiological; Process; Protein Family; Proteins; RGS Proteins; RNA; Reading; Receptor Signaling; Regulation; Research; Reverse Transcriptase Polymerase Chain Reaction; Role; Secretory Cell; Signal Transduction; Stress; Structure of beta Cell of islet; Testing; Tissues; Transplantation; Weight maintenance regimen; glucose metabolism; hormone regulation; in vitro Assay; inhibitor/antagonist; insight; insulin secretion; insulinoma; islet; knockout gene; member; neurotransmitter release; next generation sequencing; novel; programs; protein complex; protein expression; protein protein interaction; public health relevance; receptor coupling; research study; response; sensory system; stem

 DESCRIPTION (provided by applicant): Complexes of the G protein βsubunit Gβ5 with the R7 family of RGS proteins serve as GAPs for Gi family and are involved in many physiological functions. Both Gβ5 and R7 proteins were originally found in the nervous system, but recently detected in some other tissues, notably endocrine glands. The current study concentrates on the role of Gβ5-R7 in pancreatic β cells and advances an original line of research developed in this laboratory: regulation of M3 muscarinic receptor (M3R) by Gβ5-R7. The most recent study identified a novel pathway where Gβ5-R7 can potentiate M3R-induced Ca2+ influx across the plasma membrane. This effect on Ca2+ flux is consistent with the higher level of insulin secretion in the presence of Gβ5-R7. The positive effect is novel and exciting because RGS proteins are thought to be negative regulators of G protein signaling. Another puzzling detail is that M3R is a Gq-coupled receptor, and Gβ5-R7 members do not have GAP activity for Gq. The project will pursue three well-integrated aims. To determine specific β cell signaling mechanisms sensitive to Gβ5-R7, Aim 1 will target members of the M3R-to-insulin pathway with pharmacologic inhibitors or gene knockout and knockdown. Studies will be performed in pancreatic islets from WT and G 5 knockout mice and insulinoma cell line Min6. Experiments will measure insulin secretion, [Ca I] and other signaling responses to 2+ M3R agonists. Aim 2 will generate and analyze mice with β cell-specific knockout of Gβ5. Primary islets from these mice will be examined by signaling and secretion assays in vitro and long-term functionality of these islets will be studied after their intraocular transplantation into mice with induced diabete. Aim 3 will determine the effect of Gβ5-R7 complex on gene expression using a combination of RT-PCR, in situ hybridization and next generation sequencing of pancreatic islet RNA from Gβ5 KO mice. These studies will have a strong impact on understanding pancreatic β cell signaling, insulin secretion and adaptation to stress. The new mechanistic insights will provide a sustained flow of ideas for the broad area of GPCR signaling, regulation of hormone release and etiology such of metabolic disorders as diabetes and obesity.

 描述（申请人提供）：G蛋白β亚基Gβ5与RGS蛋白R7家族的复合物充当Gi家族的GAP并参与许多生理功能。 Gβ5 和 R7 蛋白最初都是在神经系统中发现的，但最近在其他一些组织中也发现了，特别是内分泌腺。目前的研究集中于 Gβ5-R7 在胰腺 β 细胞中的作用，并推进了本实验室开发的原创研究：Gβ5-R7 对 M3 毒蕈碱受体 (M3R) 的调节。最近的研究发现了一条新途径，Gβ5-R7 可以增强 M3R 诱导的 Ca2+ 跨质膜流入。这种对 Ca2+ 通量的影响与 Gβ5-R7 存在时胰岛素分泌水平较高是一致的。这种积极作用是新颖且令人兴奋的，因为 RGS 蛋白被认为是 G 蛋白信号传导的负调节因子。另一个令人费解的细节是M3R是Gq偶联受体，而Gβ5-R7成员不具有Gq的GAP活性。该项目将追求三个综合目标。为了确定对 Gβ5-R7 敏感的特定 β 细胞信号传导机制，Aim 1 将使用药理学抑制剂或基因敲除和敲除来靶向 M3R 至胰岛素途径的成员。研究将在来自 WT 和 G 5 敲除小鼠的胰岛以及胰岛素瘤细胞系 Min6 中进行。实验将测量胰岛素分泌、[Ca I] 和对 2+ M3R 激动剂的其他信号反应。目标 2 将生成并分析具有 β 细胞特异性 Gβ5 敲除的小鼠。来自这些小鼠的原代胰岛将通过体外信号传导和分泌测定进行检查，并将这些胰岛眼内移植到诱发糖尿病的小鼠体内后研究这些胰岛的长期功能。目标 3 将结合 RT-PCR、原位杂交和 Gβ5 KO 小鼠胰岛 RNA 的下一代测序，确定 Gβ5-R7 复合物对基因表达的影响。这些研究将对理解胰腺 β 细胞信号传导、胰岛素分泌和压力适应产生重大影响。新的机制见解将为 GPCR 信号传导、激素释放调节以及糖尿病和肥胖等代谢性疾病的病因学等广泛领域提供持续的思路。