Direct multiplex imaging and optogenetics of Rho GTPases enabled by near-infrared FRET

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http://hdl.handle.net/10138/303899

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Shcherbakova , D M , Cammer , N C , Huisman , T M , Verkhusha , V V & Hodgson , L 2018 , ' Direct multiplex imaging and optogenetics of Rho GTPases enabled by near-infrared FRET ' , Nature Chemical Biology , vol. 14 , no. 6 , pp. 591-+ . https://doi.org/10.1038/s41589-018-0044-1

Title: Direct multiplex imaging and optogenetics of Rho GTPases enabled by near-infrared FRET
Author: Shcherbakova, Daria M.; Cammer, Natasha Cox; Huisman, Tsipora M.; Verkhusha, Vladislav V.; Hodgson, Louis
Contributor: University of Helsinki, Medicum
Date: 2018-06
Language: eng
Number of pages: 17
Belongs to series: Nature Chemical Biology
ISSN: 1552-4450
URI: http://hdl.handle.net/10138/303899
Abstract: Direct visualization and light control of several cellular processes is a challenge, owing to the spectral overlap of available genetically encoded probes. Here we report the most red-shifted monomeric near-infrared (NIR) fluorescent protein, miRFP720, and the fully NIR Forster resonance energy transfer (FRET) pair miRFP670-miRFP720, which together enabled design of biosensors compatible with CFP-YFP imaging and blue-green optogenetic tools. We developed a NIR biosensor for Rac1 GTPase and demonstrated its use in multiplexed imaging and light control of Rho GTPase signaling pathways. Specifically, we combined the Rac1 biosensor with CFP-YFP FRET biosensors for RhoA and for Rac1-GDI binding, and concurrently used the LOV-TRAP tool for upstream Rac1 activation. We directly observed and quantified antagonism between RhoA and Rac1 dependent on the RhoA-downstream effector ROCK; showed that Rac1 activity and GDI binding closely depend on the spatiotemporal coordination between these two molecules; and simultaneously observed Rac1 activity during optogenetic manipulation of Rac1.
Subject: FLUORESCENT PROTEINS
LIVING CELLS
IN-VIVO
SPATIOTEMPORAL DYNAMICS
BACTERIAL PHYTOCHROME
CDC42 ACTIVATION
BIOSENSORS
RAC
TOOLS
COORDINATION
1182 Biochemistry, cell and molecular biology
3111 Biomedicine
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