Optogenetic interrogation reveals separable G-protein-dependent and -independent signalling linking G-protein-coupled receptors to the circadian oscillator.
Optogenetic interrogation reveals separable G-protein-dependent and -independent signalling linking G-protein-coupled receptors to the circadian oscillator.
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DOI：
10.1186/s12915-017-0380-8
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发表时间：
2017-05-15
期刊：
影响因子：
5.4
通讯作者：
Lucas RJ
中科院分区：
文献类型：
--
作者：
Bailes HJ;Milosavljevic N;Zhuang LY;Gerrard EJ;Nishiguchi T;Ozawa T;Lucas RJ
分布在哺乳动物体内的内源性昼夜节律振荡器通过各种信号分子与外部时间同步，其中一些分子与G蛋白偶联受体(GPCRs)相互作用。GPCRs可通过异源三聚体G蛋白或β阻滞素等途径调节细胞生理。我们应用光遗传学方法来确定这两种信号模式对昼夜节律的贡献。我们使用了一种光色素(JellyOp)来激活G-αS信号，比现有的替代方法具有更好的选择性和更高的灵敏度，并且这种色素的一个点突变(F112A)偏向于β-arrestin信号。当在成纤维细胞中表达时，天然的JellyOp和基于arrestin的F112A突变体都在昼夜节律时钟中驱动光依赖的阶段重置。这两种Opsins诱导的漂移在它们的昼夜相位依赖性以及它们与时钟基因诱导的关联程度上有所不同。我们的数据暗示了哺乳动物生物钟中可分离的G蛋白和arrestin输入，并建立了一对适合在活细胞中操纵GαS和βarrestin偏向信号的光遗传学工具。本文的在线版本(doi：10.1186/s12915-0170380-8)包含补充材料，授权用户可以使用。
Endogenous circadian oscillators distributed across the mammalian body are synchronised among themselves and with external time via a variety of signalling molecules, some of which interact with G-protein-coupled receptors (GPCRs). GPCRs can regulate cell physiology via pathways originating with heterotrimeric G-proteins or β-arrestins. We applied an optogenetic approach to determine the contribution of these two signalling modes on circadian phase. We employed a photopigment (JellyOp) that activates Gαs signalling with better selectivity and higher sensitivity than available alternatives, and a point mutant of this pigment (F112A) biased towards β-arrestin signalling. When expressed in fibroblasts, both native JellyOp and the F112A arrestin-biased mutant drove light-dependent phase resetting in the circadian clock. Shifts induced by the two opsins differed in their circadian phase dependence and the degree to which they were associated with clock gene induction. Our data imply separable G-protein and arrestin inputs to the mammalian circadian clock and establish a pair of optogenetic tools suitable for manipulating Gαs- and β-arrestin-biased signalling in live cells. The online version of this article (doi:10.1186/s12915-017-0380-8) contains supplementary material, which is available to authorized users.