Structure, Function and Regulation of Bombesin Receptors

铃蟾肽受体的结构、功能和调节

• 批准号: 6431974
• 负责人: James F. Battey
• 金额: --
• 依托单位: NATIONAL INSTITUTE ON DEAFNESS AND OTHER COMMUNICATION DISORDERS
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/6431974
• 关键词: G protein; autism; bombesin; gastrin releasing peptide; gene targeting; genetic regulation; guanine nucleotide binding protein; human genetic material tag; human tissue; laboratory mouse; neuropeptide receptor; phenotype; phospholipase C; phosphorylation; protein structure function; receptor coupling; site directed mutagenesis

Mammalian bombesin-like neuropeptides, gastrin releasing peptide (GRP) and neuromedin B (NMB), mediate a diverse range of biological responses in normal tissues, and stimulate the growth of some lung, pancreatic, and prostate carcinomas. Ourlaboratory cloned and characterized three structurally and pharmacologically distinct human bombesin receptors: the gastrin-releasing peptide receptor (GRP-R, or bb2); the neuromedin B receptor (NMB-R, or bb1); and bombesin receptor subtype 3 (BRS-3, or bb3). All three receptors are members of the G-protein coupled receptor superfamily, coupling to heterotrimeric G-proteins that activate phospholipase C. Several advances in our understanding of the function and regulation of these receptors has occurred within the last year: (1) we have used gene targeting strategies to create mice which lack either GRP-R or BRS-3. Mice lacking GRP-R do not exhibit bombesin-induced satiety, implicating the GRP-R as a regulator of appetite. As mice lacking GRP-R age, GRP-R targeted mice are more likely to become obese. In contrast, mice lacking BRS-3 become obese within the first few months of life.We are currently examining the hypothesis that allelic variation in the human BRS-3 gene may contribute to obesity in humans, particulary male individuals. (2) We have developed a protocol that allows us to generate uncoupled receptors embedded in a normal biological membrane. This preparation allows us to reconstitute coupling to purifiedG-proteins, examining the selectivity and enzymology of receptor-catalyzed nucleotide exchange on heterotrimeric GTP-binding proteins. Using this assay, we show that phosphorylation of serines and threonines in the C-terminal domain of the GRP-R inhibits coupling to heterotrimeric G-proteins thereby establishing that ligand-stimulated receptor phosphorylation is the switch that turns off receptor signalling. (3) We have used the yeast two-hybrid system to search for molecules that interact with the C-terminal domain, and have identified several candidates for further study, including one that has been recently shown to interact with the corresponding C-terminal domain of rhodopsin. (4) Using site-directed mutagenesis, we have shown that there are residues in the sixth transmembrane domain that are critical for selective agonist and antagonist binding, and other residues in this same domain that regulate conformational change of the GRP-R. Taken together, these studies implicate the sixth transmembrane domain as an integrator and coordinator of GRP-R function.

哺乳动物bombesin样神经肽，胃蛋白释放肽（GRP）和神经蛋白B（NMB）介导正常组织中的各种生物学反应，并刺激某些肺，胰腺和前列腺癌的生长。 我们的实验室克隆并表征了三种结构和药理学上不同的人类bombesin受体：胃蛋白释放的肽受体（GRP-R或BB2）；神经蛋白B受体（NMB-R或BB1）；和孟买受体亚型3（BRS-3或BB3）。这三个受体都是G蛋白偶联受体超家族的成员，与激活磷脂酶的异三聚体G蛋白偶联。使用基因靶向策略来创建缺乏GRP-R或BRS-3的小鼠。 缺乏GRP-R的小鼠不会表现出bombesin引起的饱腹感，这意味着Grp-R是食欲的调节剂。 由于缺乏GRP-R年龄的小鼠，GRP-R靶向小鼠更有可能肥胖。相比之下，缺乏BRS-3的小鼠在生命的头几个月内变得肥胖。我们目前正在研究以下假设：人BRS-3基因的等位基因变异可能会导致人类，特别是男性个体的肥胖。 （2）我们开发了一种协议，该方案使我们能够生成嵌入正常生物膜中的未偶联受体。 这种制备使我们能够重建耦合到纯化的蛋白质，从而检查了异源三聚体GTP结合蛋白上受体催化的核苷酸交换的选择性和酶学。 使用该测定法，我们表明，GRP-R的C末端结构域的丝氨酸和苏氨酸的磷酸化抑制与异三聚体G蛋白的耦合，从而确定配体刺激的受体磷酸化是关闭受体信号的开关。 （3）我们已经使用酵母双杂交系统来搜索与C末端结构域相互作用的分子，并确定了几个用于进一步研究的候选者，其中包括最近已证明与相应的C末端结构域相互作用的候选者Rhodopsin。 （4）使用位置定向的诱变，我们表明第六个跨膜结构域中存在残基，对于选择性激动剂和拮抗剂结合至关重要，而在同一域中的其他残基则调节了GRP-R的构象变化。综上所述，这些研究暗示了第六个跨膜结构域是GRP-R功能的集成剂和协调员。

项目成果

Molecular organization of the mouse gastrin-releasing peptide receptor gene and its promoter.

小鼠胃泌素释放肽受体基因及其启动子的分子结构。

• DOI: 10.1016/s0378-1119(99)00563-6
• 发表时间: 2000
• 期刊: Gene
• 影响因子: 3.5
• 作者: Weber,HC;Jensen,RT;Battey,JF
• 通讯作者: Battey,JF

Two amino acids in the sixth transmembrane segment of the mouse gastrin-releasing peptide receptor are important for receptor activation.

小鼠胃泌素释放肽受体第六跨膜片段中的两个氨基酸对于受体激活很重要。

• 发表时间: 2000
• 期刊: The Journal of pharmacology and experimental therapeutics
• 作者: Lin,Y;Jian,X;Lin,Z;Kroog,GS;Mantey,S;Jensen,RT;Battey,J;Northup,J
• 通讯作者: Northup,J

Comparative pharmacology of the nonpeptide neuromedin B receptor antagonist PD 168368.

非肽神经调节肽 B 受体拮抗剂 PD 168368 的药理学比较。

• 发表时间: 1999
• 期刊: The Journal of pharmacology and experimental therapeutics
• 作者: Ryan,RR;Katsuno,T;Mantey,SA;Pradhan,TK;Weber,HC;Coy,DH;Battey,JF;Jensen,RT
• 通讯作者: Jensen,RT