Optogenetic interrogation reveals separable G-protein-dependent and -independent signalling linking G-protein-coupled receptors to the circadian oscillator.

BMC Biology
Helena J BailesRobert J Lucas

Abstract

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.

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Citations

May 26, 2018·Proceedings of the National Academy of Sciences of the United States of America·Elliot GerrardRobert J Lucas

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Datasets Mentioned

BETA
AB435549

Methods Mentioned

BETA
transfect
ELISA
transfection
proximity ligation
FRET
Assay
enzyme-linked immunoassay

Software Mentioned

JellyOp
Opto
GraphPad
Clockwise
MetaVue
Excel
Lumicycle
Softworx
per2
GraphPad Prism

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