Orthogonal fluorescent chemogenetic reporters for multicolor imaging

TY - JOUR

T1 - Orthogonal fluorescent chemogenetic reporters for multicolor imaging

AU - Tebo, Alison G.

AU - Moeyaert, Benjamien

AU - Thauvin, Marion

AU - Carlon-Andres, Irene

AU - Böken, Dorothea

AU - Volovitch, Michel

AU - Padilla-Parra, Sergi

AU - Dedecker, Peter

AU - Vriz, Sophie

AU - Gautier, Arnaud

N1 - Funding Information: We thank K.D. Wittrup, for providing us with the pCTCON2 vector and the EBY100 yeast strain for the yeast display selection. We also thank the flow cytometry facility Cytométrie Imagerie Saint-Antoine of UMS LUMIC at the Faculty of Medicine of Sorbonne University and, particularly, A. Munier for her assistance. This work has been supported by the European Research Council (ERC-2016-CoG-724705 FLUOSWITCH to A.G., ERC StG 714688 NanoCellActivity to P.D. and ERC-2019-CoG-863869 FUSION to S.P.-P.), the Wellcome Trust Core Award (203141 to S.P.-P.), and the Research Foundation-Flanders (G0B8817N to P.D. and 1514319N to B.M.). Publisher Copyright: © 2020, The Author(s), under exclusive licence to Springer Nature America, Inc. Copyright: Copyright 2021 Elsevier B.V., All rights reserved.

PY - 2021/1

Y1 - 2021/1

N2 - Spectrally separated fluorophores allow the observation of multiple targets simultaneously inside living cells, leading to a deeper understanding of the molecular interplay that regulates cell function and fate. Chemogenetic systems combining a tag and a synthetic fluorophore provide certain advantages over fluorescent proteins since there is no requirement for chromophore maturation. Here, we present the engineering of a set of spectrally orthogonal fluorogen-activating tags based on the fluorescence-activating and absorption shifting tag (FAST) that are compatible with two-color, live-cell imaging. The resulting tags, greenFAST and redFAST, demonstrate orthogonality not only in their fluorogen recognition capabilities, but also in their one- and two-photon absorption profiles. This pair of orthogonal tags allowed the creation of a two-color cell cycle sensor capable of detecting very short, early cell cycles in zebrafish development and the development of split complementation systems capable of detecting multiple protein–protein interactions by live-cell fluorescence microscopy. [Figure not available: see fulltext.].

AB - Spectrally separated fluorophores allow the observation of multiple targets simultaneously inside living cells, leading to a deeper understanding of the molecular interplay that regulates cell function and fate. Chemogenetic systems combining a tag and a synthetic fluorophore provide certain advantages over fluorescent proteins since there is no requirement for chromophore maturation. Here, we present the engineering of a set of spectrally orthogonal fluorogen-activating tags based on the fluorescence-activating and absorption shifting tag (FAST) that are compatible with two-color, live-cell imaging. The resulting tags, greenFAST and redFAST, demonstrate orthogonality not only in their fluorogen recognition capabilities, but also in their one- and two-photon absorption profiles. This pair of orthogonal tags allowed the creation of a two-color cell cycle sensor capable of detecting very short, early cell cycles in zebrafish development and the development of split complementation systems capable of detecting multiple protein–protein interactions by live-cell fluorescence microscopy. [Figure not available: see fulltext.].

UR - http://www.scopus.com/inward/record.url?scp=85089260101&partnerID=8YFLogxK

U2 - 10.1038/s41589-020-0611-0

DO - 10.1038/s41589-020-0611-0

M3 - Article

AN - SCOPUS:85089260101

SN - 1552-4450

VL - 17

SP - 30

EP - 38

JO - Nature Chemical Biology

JF - Nature Chemical Biology

IS - 1

ER -