INTERACTIONS BETWEEN GABA B AND CA2+-SENSING RECEPTORS

GABA B 和 CA2 传感受体之间的相互作用

• 批准号: 7186125
• 负责人: Wenhan Chang
• 金额: $ 24.83万
• 依托单位: NORTHERN CALIFORNIA INSTITUTE/RES/EDU
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-03-06 至 2009-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7186125
• 关键词: Address; Affect; Aminobutyric Acid; Aminobutyric Acids; Antibodies; Anxiety; Apoptosis; Attention; Biochemical; Body Size; Bone Growth; Bone and Cartilage Funding; Brain; Calcium-Sensing Receptors; Cartilage; Cell Proliferation; Cell physiology; Cells; Chondrocytes; Chromosome Pairing; Co-Immunoprecipitations; Complex; Coupling; Differentiation Antigens; Differentiation and Growth; Distal; Doctor of Philosophy; Epilepsy; Epiphysial cartilage; Equilibrium; Excretory function; Family; Functional disorder; Future; G-Protein-Coupled Receptors; GABA Receptor; GABA-B Receptor; Genes; Glutamate Receptor; Height; Human; Hyperactive behavior; In Vitro; Intestines; Kidney; Knock-out; Ligand Binding; Ligands; Link; Localized; Membrane; Mental Depression; Metabotropic Glutamate Receptors; Minerals; Molecular Conformation; Morphology; Mus; Nervous System Physiology; Neuraxis; Neurons; Numbers; Osteogenesis; Pain; Parathyroid Hormones; Parathyroid gland; Phenotype; Pheromone Receptors; Phospholipase C; Physiological; Physiology; Play; Population; Process; Property; Proteins; RRM1 gene; Receptor Mediated Signal Transduction; Receptor Signaling; Regulation; Research Personnel; Role; Schizophrenia; Signal Transduction; Sleep Disorders; Spinal cord injury; Stress; Synapses; System; Testing; Thinking; Tissues; Work; addiction; base; brain tissue; cartilage development; cognitive function; extracellular; gamma-Aminobutyric Acid; human PTH protein; in vivo; insight; member; monomer; programs; receptor; receptor expression; receptor function; response; subunit 1 GABA type B receptor; trafficking

DESCRIPTION (provided by applicant): Family C receptors of the G-protein coupled receptor superfamily include metabotropic ?-aminobutyric acid (GABA)-B receptors (GABA-B-Rs), metabotropic glutamate receptors (mGluRs), and Ca2+-sensing receptors (CaRs). These receptors function in dimeric or multimeric forms which maintain the proper conformations for ligand binding, trafficking, and coupling to downstream effectors. A functional GABA-B-R consists of a heterodimer between the GABA-B-R1 and R2, two distinct gene products. CaRs function as homodimers in target cells (i.e., parathyroid and kidney), and they also can heterodimerize with mGluRs in the brain. We recently found that CaRs can also form heteromeric complexes with GABA-B-R 1s in HEK-293 cells transfected with both receptor cDNAs. Co-expressing GABA-B-R1s along with CaRs suppressed CaR protein levels and the responsiveness of phospholipase C activation in the cells to changes in the extracellular [Ca2+] ([Ca2+]e), thus supporting the idea that GABA-B-R1s alter CaR expression and function via a heteromeric receptor complex. Whether such co-associations occur in the target cells in vivo has received little attention. Both CaRs and GABA-B-Rs are present in many tissues - brain, kidney, parathyroid, intestine, bone and cartilage. In cultured mouse growth plate chondrocytes (mGPCs), knocking out the GABA-B-R1 genes dramatically increased the level of CaR protein and signaling responses to high [Ca2+]e, supporting a role for GABA-B-R1s in regulating the expression and function of the CaRs in mGPCs. Our hypothesis is that GABA-B-R1s modulate the expression and signal transduction of CaRs by the formation of heteromeric complexes between the two molecules and that this interaction impacts on the signaling responses and function of target cells. To test this hypothesis, we propose the following aims. AIM 1: To determine whether CaRs and GABA-B-R1s interact to modulate CaR expression and signal transduction. We will address this by (a) co-expressing GABA-B-R1s with CaRs in HEK-293 cells and assessing whether their association affects the synthesis and or degradation of CaR protein and alters the signal transduction properties of CaRs; (b) testing the effects of blocking GABA-B-R1 expression in mGPCs on the synthesis and or degradation of CaR protein and on CaR-mediated signal transduction; (c) ascertaining whether GABA-B-R1s co-associate with CaRs in mGPCs; and (d) examining the effects of blocking GABA-B-R1 expression on the proliferation, apoptosis, and expression of markers of differentiation -- processes responsive to changes in the [Ca2+]e in mGPCs. AIM 2: To determine the impact of the GABA- B-R1 in cartilage in vivo by examining the effect of cartilage-specific knockout of the GABA-B-R1 genes on CaR expression, morphology, cell proliferation, apoptosis, and the expression of markers of differentiation in growth plate cartilage in mice. Completion of this work will provide insights into how CaRs and GABA-B-R1s interact in a physiological context and how such an interaction influences cartilage development. CaRs control mineral balance in the body, and GABA-B-Rs are essential to nerve cell function. The growth plate provides the cells and proteins that initiate new bone formation, which ultimately determines our height and body size. This work can open up future studies targeted to developing therapies to modulate the function of CaRs, GABA-B-Rs, or the heteromeric complexes of these two receptors.

描述(申请人提供)：G蛋白偶联受体超家族的C家族受体包括代谢性β-氨基丁酸(GABA)-B受体(GABA-B-Rs)、代谢性谷氨酸受体(MGluRs)和钙敏感受体(CARS)。这些受体以二聚体或多聚体的形式发挥作用，为配体的结合、运输和与下游效应器的偶联保持适当的构象。一个功能性的GABA-B-R由两个不同的基因产物--GABA-B-R1和R2之间的异源二聚体组成。CARS在靶细胞(即甲状旁腺和肾脏)中作为同源二聚体发挥作用，它们也可以与大脑中的mGluRs异二聚体。我们最近发现，在HEK-293细胞中，CARS还可以与GABA-B-R1S形成异构体复合体。与CARS共同表达的GABA-B-R1s抑制了细胞内CAR蛋白水平和磷脂酶C激活对细胞外[Ca2+]e变化的反应，从而支持了GABA-B-R1s通过异构体受体复合体改变CAR表达和功能的观点。在体内的靶细胞中是否存在这样的共同联系，人们几乎没有注意到。CARS和GABA-B受体都存在于许多组织中--脑、肾、甲状旁腺、肠、骨和软骨。在培养的小鼠生长板软骨细胞中，敲除GABA-B-R1基因显著增加了CAR蛋白水平和对高[Ca~(2+)]e的信号反应，支持了GABA-B-R1s在调节MGPC中CARS的表达和功能中的作用。我们的假设是，GABA-B-R1s通过在两个分子之间形成异构体复合体来调节CARS的表达和信号转导，这种相互作用影响靶细胞的信号反应和功能。为了验证这一假设，我们提出了以下目标。目的1：确定CARS和GABA-B-R1s是否相互作用调节CAR的表达和信号转导。我们将通过以下方式解决这一问题：(A)在HEK-293细胞中与CARS共表达GABA-B-R1s，并评估它们的关联是否影响CAR蛋白的合成和/或降解并改变CARS的信号转导特性；(B)检测阻断mGPC中GABA-B-R1s的表达对CAR蛋白的合成和/或降解以及CAR介导的信号转导的影响；(C)确定GABA-B-R1s是否与mGPC中的CARS共同作用；以及(D)检测阻断GABA-B-R1表达对MGPC增殖、凋亡和分化标记物表达的影响--这些过程是对mGPC中[Ca~(2+)]e变化的反应。目的：通过检测软骨特异性敲除GABA-B-R1基因对小鼠生长板软骨CAR表达、细胞形态、细胞增殖、细胞凋亡及分化标志物表达的影响，探讨GABA-B-R1基因在体内对软骨的影响。这项工作的完成将提供对CARS和GABA-B-R1s在生理环境中如何相互作用以及这种相互作用如何影响软骨发育的见解。CARS控制着体内的矿物质平衡，而GABA-B-Rs是神经细胞功能所必需的。生长板提供启动新骨形成的细胞和蛋白质，这最终决定了我们的身高和身体尺寸。这项工作可能会开启未来的研究，目标是开发治疗方法来调节CARS、GABA-B-Rs或这两种受体的异构体复合体的功能。