title: Probing plant membranes with FM dyes: tracking, dragging or blocking?
abstract: Remarkable progress in various techniques of in vivo fluorescence microscopy has brought an urgent need for reliable markers for tracking cellular structures and processes. The goal of this manuscript is to describe unexplored effects of the FM (Fei Mao) styryl dyes, which are widely used probes that label processes of endocytosis and vesicle trafficking in eukaryotic cells. Although there are few reports on the effect of styryl dyes on membrane fluidity and the activity of mammalian receptors, FM dyes have been considered as reliable tools for tracking of plant endocytosis. Using plasma membrane-localized transporters for the plant hormone auxin in tobacco BY-2 and Arabidopsis thaliana cell suspensions, we show that routinely used concentrations of FM 4-64 and FM 5-95 trigger transient re-localization of these proteins, and FM 1-43 affects their activity. The active process of re-localization is blocked neither by inhibitors of endocytosis nor by cytoskeletal drugs. It does not occur in A. thaliana roots and depends on the degree of hydrophobicity (lipophilicity) of a particular FM dye. Our results emphasize the need for circumspection during in vivo studies of membrane proteins performed using simultaneous labelling with FM dyes.
authors.affiliation: Institute of Experimental Botany, Academy of Sciences of the Czech Republic, Rozvojová 263, 165 02 Prague 6, Czech Republic.
authors.forename: Adriana
authors.initials: A
authors.lastname: Jelínková
authors.forename: Katerina
authors.initials: K
authors.lastname: Malínská
authors.forename: Sibu
authors.initials: S
authors.lastname: Simon
authors.forename: Jürgen
authors.initials: J
authors.lastname: Kleine-Vehn
authors.forename: Markéta
authors.initials: M
authors.lastname: Parezová
authors.forename: Premysl
authors.initials: P
authors.lastname: Pejchar
authors.forename: Martin
authors.initials: M
authors.lastname: Kubes
authors.forename: Jan
authors.initials: J
authors.lastname: Martinec
authors.forename: Jirí
authors.initials: J
authors.lastname: Friml
authors.forename: Eva
authors.initials: E
authors.lastname: Zazímalová
authors.forename: Jan
authors.initials: J
authors.lastname: Petrásek
mesh_terms: D017360:Arabidopsis / Q000166:cytology; D002460:Cell Line; D002462:Cell Membrane / Q000737:chemistry*; D004705:Endocytosis; D005456:Fluorescent Dyes / Q000494:pharmacology*; D007210:Indoleacetic Acids / Q000378:metabolism; D010940:Plant Proteins / Q000187:drug effects*; D021381:Protein Transport; D011726:Pyridinium Compounds / Q000494:pharmacology; D000644:Quaternary Ammonium Compounds / Q000494:pharmacology
chemical_list: C092350:FM 4-64; D005456:Fluorescent Dyes; D007210:Indoleacetic Acids; C508379:N-(3-trimethylammoniumpropyl)-4-(6-(4-(diethylamino)phenyl)hexatrienyl)pyridinium; D010940:Plant Proteins; D011726:Pyridinium Compounds; D000644:Quaternary Ammonium Compounds