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Nature:构象生物传感器揭示内涵体上的G蛋白偶联受体信号

G蛋白偶联受体

G蛋白偶联受体(G Protein-Coupled Receptors, GPCRs),是一大类膜蛋白受体的统称。这类受体的共同点是其立体结构中都有七个跨膜α螺旋,且其肽链的C端和连接第5和第6个跨膜螺旋的胞内环上都有G蛋白(鸟苷酸结合蛋白)的结合位点。目前为止,研究显示G蛋白偶联受体只见于真核生物之中,而且参与了很多细胞信号转导过程。在这些过程中,G蛋白偶联受体能结合细胞周围环境中的化学物质并激活细胞内的一系列信号通路,最终引起细胞状态的改变。

不管是长期以来科学家认为:G蛋白偶联受体(GPCR)信号转导常发生在质膜上,还是后来提出的G蛋白偶联受体内化受到G蛋白依赖机制的限制,人们始终无法找到直接的证据证实这些假设。现在,美国加州大学、密歇根大学等机构的研究人员终于证实了内涵体上GPCR的激活。相关研究论文刊登在近期出版的《自然》(Nature)杂志上。

长期以来,分子药理学上有一个理论旨就是由G蛋白偶联受体(GPCR)介导的典型信号转导仅局限于质膜上。这种传统观点是基于有限的分析方法以及无亚细胞分辨率条件下。

随后,人们提出了G蛋白偶联受体(GPCRs)内化受到G蛋白依赖机制的限制,如:G蛋白偶联受体(GPCR)的激活引起持续性G蛋白的离散形式或G蛋白偶联受体(GPCRs)内化确实可以有助于G蛋白介导的应激反应。

支持后来那些假设的证据往往是间接的或者需要服从于另外一种解释。如果GPCRs受体内化发生在内涵体上,且呈现活性状态,那现在就不清楚了。

美国加州大学一个研究小组描述了特异构象单域抗体(也称纳米抗体)的应用,可直接证实活体哺乳动物细胞中β 2 -肾上腺素的激活,它是一个典型的G蛋白偶联受体(GPCR),其同源蛋白G s。

此项研究发现,肾上腺素受体激动剂异丙肾上腺素如预期的一样能促进质膜和早期内涵体膜上受体与G蛋白的激活。与此同时,激动剂应用后的几分钟内,内化受体也为细胞内总cAMP水平做出贡献。

这些发现给内涵体和质膜上典型GPCR信号的假设提供了直接证据,为探讨体外一动态构象变化提供了一个通用策略。

了解更多:

Conformational biosensors reveal GPCR signalling from endosomes

Nature, 20 March 2013 | doi:10.1038/nature12000

A long-held tenet of molecular pharmacology is that canonical signal transduction mediated by G-protein-coupled receptor (GPCR) coupling to heterotrimeric G proteins is confined to the plasma membrane. Evidence supporting this traditional view is based on analytical methods that provide limited or no subcellular resolution. It has been subsequently proposed that signalling by internalized GPCRs is restricted to G-protein-independent mechanisms such as scaffolding by arrestins, or GPCR activation elicits a discrete form of persistent G protein signalling, or that internalized GPCRs can indeed contribute to the acute G-protein-mediated response. Evidence supporting these various latter hypotheses is indirect or subject to alternative interpretation, and it remains unknown if endosome-localized GPCRs are even present in an active form. Here we describe the application of conformation-specific single-domain antibodies (nanobodies) to directly probe activation of the β2-adrenoceptor, a prototypical GPCR, and its cognate G protein, Gs , in living mammalian cells. We show that the adrenergic agonist isoprenaline promotes receptor and G protein activation in the plasma membrane as expected, but also in the early endosome membrane, and that internalized receptors contribute to the overall cellular cyclic AMP response within several minutes after agonist application. These findings provide direct support for the hypothesis that canonical GPCR signalling occurs from endosomes as well as the plasma membrane, and suggest a versatile strategy for probing dynamic conformational change in vivo.

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