title: Small fluorescence-activating and absorption-shifting tag for tunable protein imaging in vivo.
abstract: This paper presents Yellow Fluorescence-Activating and absorption-Shifting Tag (Y-FAST), a small monomeric protein tag, half as large as the green fluorescent protein, enabling fluorescent labeling of proteins in a reversible and specific manner through the reversible binding and activation of a cell-permeant and nontoxic fluorogenic ligand (a so-called fluorogen). A unique fluorogen activation mechanism based on two spectroscopic changes, increase of fluorescence quantum yield and absorption red shift, provides high labeling selectivity. Y-FAST was engineered from the 14-kDa photoactive yellow protein by directed evolution using yeast display and fluorescence-activated cell sorting. Y-FAST is as bright as common fluorescent proteins, exhibits good photostability, and allows the efficient labeling of proteins in various organelles and hosts. Upon fluorogen binding, fluorescence appears instantaneously, allowing monitoring of rapid processes in near real time. Y-FAST distinguishes itself from other tagging systems because the fluorogen binding is highly dynamic and fully reversible, which enables rapid labeling and unlabeling of proteins by addition and withdrawal of the fluorogen, opening new exciting prospects for the development of multiplexing imaging protocols based on sequential labeling.
authors.affiliation: Department of Chemistry, École Normale Supérieure, Paris Sciences et Lettres (PSL) Research University, F-75005 Paris, France; Sorbonne Universités, Université Pierre et Marie Curie, UMR 8640 PASTEUR, F-75005 Paris, France; CNRS, UMR 8640 PASTEUR, F-75005 Paris, France;
authors.forename: Marie-Aude
authors.initials: MA
authors.lastname: Plamont
authors.affiliation: Department of Chemistry, École Normale Supérieure, Paris Sciences et Lettres (PSL) Research University, F-75005 Paris, France; Sorbonne Universités, Université Pierre et Marie Curie, UMR 8640 PASTEUR, F-75005 Paris, France; CNRS, UMR 8640 PASTEUR, F-75005 Paris, France;
authors.forename: Emmanuelle
authors.initials: E
authors.lastname: Billon-Denis
authors.affiliation: Department of Chemistry, École Normale Supérieure, Paris Sciences et Lettres (PSL) Research University, F-75005 Paris, France; Sorbonne Universités, Université Pierre et Marie Curie, UMR 8640 PASTEUR, F-75005 Paris, France; CNRS, UMR 8640 PASTEUR, F-75005 Paris, France;
authors.forename: Sylvie
authors.initials: S
authors.lastname: Maurin
authors.affiliation: Centre for Interdisciplinary Research in Biology, CNRS UMR 7241, INSERM U1050, Labex MemoLife, Paris Sciences et Lettres (PSL) Research University/Collège de France, F-75005 Paris, France;
authors.forename: Carole
authors.initials: C
authors.lastname: Gauron
authors.affiliation: Department of Chemistry, École Normale Supérieure, Paris Sciences et Lettres (PSL) Research University, F-75005 Paris, France; Sorbonne Universités, Université Pierre et Marie Curie, UMR 8640 PASTEUR, F-75005 Paris, France; CNRS, UMR 8640 PASTEUR, F-75005 Paris, France;
authors.forename: Frederico M
authors.initials: FM
authors.lastname: Pimenta
authors.affiliation: École Normale Supérieure, Institut de Biologie de l'École Normale Supérieure (IBENS), CNRS UMR 8197, INSERM U1024, Paris Sciences et Lettres (PSL) Research University, F-75005 Paris, France;
authors.forename: Christian G
authors.initials: CG
authors.lastname: Specht
authors.affiliation: Department of Chemistry, École Normale Supérieure, Paris Sciences et Lettres (PSL) Research University, F-75005 Paris, France; Sorbonne Universités, Université Pierre et Marie Curie, UMR 8640 PASTEUR, F-75005 Paris, France; CNRS, UMR 8640 PASTEUR, F-75005 Paris, France;
authors.forename: Jian
authors.initials: J
authors.lastname: Shi
authors.affiliation: Department of Chemistry, École Normale Supérieure, Paris Sciences et Lettres (PSL) Research University, F-75005 Paris, France; Sorbonne Universités, Université Pierre et Marie Curie, UMR 8640 PASTEUR, F-75005 Paris, France; CNRS, UMR 8640 PASTEUR, F-75005 Paris, France;
authors.forename: Jérôme
authors.initials: J
authors.lastname: Quérard
authors.affiliation: Department of Chemistry, École Normale Supérieure, Paris Sciences et Lettres (PSL) Research University, F-75005 Paris, France; Sorbonne Universités, Université Pierre et Marie Curie, UMR 8640 PASTEUR, F-75005 Paris, France; CNRS, UMR 8640 PASTEUR, F-75005 Paris, France;
authors.forename: Buyan
authors.initials: B
authors.lastname: Pan
authors.affiliation: Department of Chemistry, École Normale Supérieure, Paris Sciences et Lettres (PSL) Research University, F-75005 Paris, France; Sorbonne Universités, Université Pierre et Marie Curie, UMR 8640 PASTEUR, F-75005 Paris, France; CNRS, UMR 8640 PASTEUR, F-75005 Paris, France;
authors.forename: Julien
authors.initials: J
authors.lastname: Rossignol
authors.affiliation: Laboratoire de Biologie Physico-Chimique des Protéines Membranaires, Institut de Biologie Physico-Chimique, CNRS UMR 7099, Université Paris Diderot, Sorbonne Paris Cité, Paris Sciences et Lettres (PSL) Research  University, F-75005 Paris, France.
authors.forename: Karine
authors.initials: K
authors.lastname: Moncoq
authors.affiliation: Institut de Chimie des Substances Naturelles, CNRS UPR 2301, Université Paris-Saclay, 91198 Gif-sur-Yvette, France;
authors.forename: Nelly
authors.initials: N
authors.lastname: Morellet
authors.affiliation: Centre for Interdisciplinary Research in Biology, CNRS UMR 7241, INSERM U1050, Labex MemoLife, Paris Sciences et Lettres (PSL) Research University/Collège de France, F-75005 Paris, France; École Normale Supérieure, Institut de Biologie de l'École Normale Supérieure (IBENS), CNRS UMR 8197, INSERM U1024, Paris Sciences et Lettres (PSL) Research University, F-75005 Paris, France;
authors.forename: Michel
authors.initials: M
authors.lastname: Volovitch
authors.affiliation: Institut de Chimie des Substances Naturelles, CNRS UPR 2301, Université Paris-Saclay, 91198 Gif-sur-Yvette, France;
authors.forename: Ewen
authors.initials: E
authors.lastname: Lescop
authors.affiliation: Department of Chemistry, École Normale Supérieure, Paris Sciences et Lettres (PSL) Research University, F-75005 Paris, France; Sorbonne Universités, Université Pierre et Marie Curie, UMR 8640 PASTEUR, F-75005 Paris, France; CNRS, UMR 8640 PASTEUR, F-75005 Paris, France;
authors.forename: Yong
authors.initials: Y
authors.lastname: Chen
authors.affiliation: École Normale Supérieure, Institut de Biologie de l'École Normale Supérieure (IBENS), CNRS UMR 8197, INSERM U1024, Paris Sciences et Lettres (PSL) Research University, F-75005 Paris, France;
authors.forename: Antoine
authors.initials: A
authors.lastname: Triller
authors.affiliation: Centre for Interdisciplinary Research in Biology, CNRS UMR 7241, INSERM U1050, Labex MemoLife, Paris Sciences et Lettres (PSL) Research University/Collège de France, F-75005 Paris, France; Université Paris Diderot Sorbonne Paris Cité, 75205 Paris, France.
authors.forename: Sophie
authors.initials: S
authors.lastname: Vriz
authors.affiliation: Department of Chemistry, École Normale Supérieure, Paris Sciences et Lettres (PSL) Research University, F-75005 Paris, France; Sorbonne Universités, Université Pierre et Marie Curie, UMR 8640 PASTEUR, F-75005 Paris, France; CNRS, UMR 8640 PASTEUR, F-75005 Paris, France;
authors.forename: Thomas
authors.initials: T
authors.lastname: Le Saux
authors.affiliation: Department of Chemistry, École Normale Supérieure, Paris Sciences et Lettres (PSL) Research University, F-75005 Paris, France; Sorbonne Universités, Université Pierre et Marie Curie, UMR 8640 PASTEUR, F-75005 Paris, France; CNRS, UMR 8640 PASTEUR, F-75005 Paris, France; arnaud.gautier@ens.fr ludovic.jullien@ens.fr.
authors.forename: Ludovic
authors.initials: L
authors.lastname: Jullien
authors.affiliation: Department of Chemistry, École Normale Supérieure, Paris Sciences et Lettres (PSL) Research University, F-75005 Paris, France; Sorbonne Universités, Université Pierre et Marie Curie, UMR 8640 PASTEUR, F-75005 Paris, France; CNRS, UMR 8640 PASTEUR, F-75005 Paris, France; arnaud.gautier@ens.fr ludovic.jullien@ens.fr.
authors.forename: Arnaud
authors.initials: A
authors.lastname: Gautier
mesh_terms: D065638:Absorption, Radiation; D000818:Animals; D002470:Cell Survival; D004625:Embryo, Nonmammalian / Q000378:metabolism; D005453:Fluorescence; D005818:Genetic Engineering; D006367:HeLa Cells; D006801:Humans; D021621:Imaging, Three-Dimensional; D011993:Recombinant Fusion Proteins / Q000378:metabolism*; D013194:Staining and Labeling; D013347:Subcellular Fractions / Q000378:metabolism; D015027:Zebrafish / Q000196:embryology
chemical_list: D011993:Recombinant Fusion Proteins
keywords: directed evolution; fluorescence imaging; fluorogenic ligand