Analysis of the molecular mechanisms of formation and trafficking to the cell surface of the G protein-coupled receptors that require accessory proteins for their expression.

分析需要辅助蛋白表达的 G 蛋白偶联受体形成和运输到细胞表面的分子机制。

• 批准号: 13680846
• 负责人: UEZONO Yasuhito
• 金额: $ 2.3万
• 依托单位: Nagasaki University Graduate School of Biomedical Sciences
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2001
• 资助国家: 日本
• 起止时间: 2001 至 2002
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-13680846/
• 关键词: GABA_B receptors; G-protein coupled receptors; Heteromultimerization; Xenopus oocytes; Heterologous expression; FRET; Electrophysiology; Laser confocal microscopy; 内向き整流性Kチャネル; CFTR

γ-Amino butyric acids (GABA) is known to act as an inhibitory neurotransmitter on the central and peripheral nervous system. Receptors for GABA are divided into two types : one belongs to ionotropic receptors (GABA_A and GABA_C) and another belongs to metabotoropic G-protein coupled receptors (GABA_B). GABA_B receptor has been cloned in 1997 as a GABA_<B1> receptor and since then, many of works have been focused on the heterologous functional expression of the cloned GABA_B receptors. However, none of the expression study has succeeded. Accordingly the failure of the expression study suggested us the existence of some additional protein(s) for the functional expression of GABA_B receptors. Now functional GABA_B receptors are known to be required to be formed heterodimer with GABA_<B1> and GBA_<B2> receptors.In the present study we focused on how functional GABA_B receptors are constructed as heterodimer and how the receptors are trafficked to the cell membrane, by the use of multiple assay systems. We found that functional GABA_B receptors are already formed before trafficked to cell membranes. We also found that GABA_<B1> receptors are required for their ligand binding while GABA_<B2> receptors are indispensable for their intracellular signaling through heteromeric G proteins.We are still in progress of our study and hope to continue to clarify the mechanism how GABA_B receptors are formed and why GABA_B receptors are required to be heteromultimer for their functional expression.

已知γ-氨基丁酸（GABA）作为中枢和外周神经系统的抑制性神经递质。GABA受体分为两类：一类是离子型受体（GABA_A和GABA_C），另一类是代谢型G蛋白偶联受体（GABA_B）。GABA_B受体是1997年克隆的一种GABA_<B1>受体，从那时起，许多工作都集中在克隆的GABA_B受体的异源功能表达上。然而，没有一个表达研究是成功的。因此，表达研究的失败提示我们存在一些额外的蛋白质用于GABA_B受体的功能表达。目前已知功能性GABA_（B）受体需要与GABA_（1）和GABA_（2）受体形成异源二聚体<B1><B2>，本研究利用多种分析系统，着重探讨功能性GABA_（B）受体是如何形成异源二聚体的，以及受体是如何转运到细胞膜上的。我们发现功能性GABA_B受体在进入细胞膜之前就已经形成。我们还发现GABA_B<B1>受体是其配体结合所必需的，而GABA_B<B2>受体是其通过异聚体G蛋白进行细胞内信号转导所必需的，我们的研究仍在进行中，希望能继续阐明GABA_B受体的形成机制以及GABA_B受体功能表达需要异聚体的原因。

项目成果

Shiraishi, M., Shibuya, I., Uezono, Y., et al.: "A neurosteroid anesthetic, alphaxalone, inhibits nicotinic acetylcholine receptors in cultured bovine adrenal chromaffin cells"Anesthesia and Analgesia. 95. 900-906 (2002)

Shiraishi, M.、Shibuya, I.、Uezono, Y. 等人：“神经类固醇麻醉剂阿法沙酮可抑制培养的牛肾上腺嗜铬细胞中的烟碱乙酰胆碱受体”麻醉和镇痛。

NAGAOKA, E., MINAMI, K., SHIGA, Y., UEZONO, Y., SHIRAISHI, M., AOYAMA, K., SHIGEMATSU, A.: "Tramadol has no effect on renal blood flow - despite increased serum norepinephrine levels - in anesthetized rats : Implication for analgesia in renal insufficienc

NAGAOKA, E.、MINAMI, K.、SHIGA, Y.、UEZONO, Y.、SHIRAISHI, M.、AOYAMA, K.、SHIGEMATSU, A.：“曲马多对肾血流量没有影响 - 尽管血清去甲肾上腺素水平升高

TSUDA, Y., OKAZAKI, M., UEZONO, Y., OSAJIMA, A., KATO, H., OKUDA, H., OISHI, Y., YASHIRO, A., NAKASHIMA, Y.: "Activation of extracellular signal-regulated kinases is essential for pure transmural pressure-induced proliferation of vascular smooth muscle ce

TSUDA, Y., OKAZAKI, M., UEZONO, Y., OSAJIMA, A., KATO, H., OKUDA, H., OISHI, Y., YASHIRO, A., NAKASHIMA, Y.：“细胞外信号的激活

Shiraishi, S., Yamamoto, R., Uezono, Y., et al.: "Down-regulation of cell surface insulin receptors by sarco(endo)plasmic reticulum Ca2+-ATPase inhibitor in adrenal chromaffin cells"Brain Research. 898. 152-157 (2001)

Shiraishi, S.、Yamamoto, R.、Uezono, Y. 等人：“肾上腺嗜铬细胞中肌（内）质网 Ca2 -ATP 酶抑制剂下调细胞表面胰岛素受体”脑研究。

Kaibara, M., Nagase, Y., Uezono, Y., et al.: "GTP-γS-induced Ca2+-activated Cl-currents : its stable induction by Gqα overexpression in Xenopus oocytes"Japanese Journal of Pharmacology. 86. 244-247 (2001)

Kaibara, M.、Nagase, Y.、Uezono, Y. 等人：“GTP-γS 诱导的 Ca2+ 激活的 Cl 电流：爪蟾卵母细胞中 Gqα 过表达的稳定诱导”《日本药理学杂志》86. 244。 -247 (2001)