title: Fluorescent castasterone reveals BRI1 signaling from the plasma membrane.
abstract: Receptor-mediated endocytosis is an integral part of signal transduction as it mediates signal attenuation and provides spatial and temporal dimensions to signaling events. One of the best-studied leucine-rich repeat receptor-like kinases in plants, BRASSINOSTEROID INSENSITIVE 1 (BRI1), perceives its ligand, the brassinosteroid (BR) hormone, at the cell surface and is constitutively endocytosed. However, the importance of endocytosis for BR signaling remains unclear. Here we developed a bioactive, fluorescent BR analog, Alexa Fluor 647-castasterone (AFCS), and visualized the endocytosis of BRI1-AFCS complexes in living Arabidopsis thaliana cells. Impairment of endocytosis dependent on clathrin and the guanine nucleotide exchange factor for ARF GTPases (ARF-GEF) GNOM enhanced BR signaling by retaining active BRI1-ligand complexes at the plasma membrane. Increasing the trans-Golgi network/early endosome pool of BRI1-BR complexes did not affect BR signaling. Our findings provide what is to our knowledge the first visualization of receptor-ligand complexes in plants and reveal clathrin- and ARF-GEF-dependent endocytic regulation of BR signaling from the plasma membrane.
authors.affiliation: Department of Plant Systems Biology, VIB, Ghent, Belgium.
authors.forename: Niloufer G
authors.initials: NG
authors.lastname: Irani
authors.forename: Simone
authors.initials: S
authors.lastname: Di Rubbo
authors.forename: Evelien
authors.initials: E
authors.lastname: Mylle
authors.forename: Jos
authors.initials: J
authors.lastname: Van den Begin
authors.forename: Joanna
authors.initials: J
authors.lastname: Schneider-Pizoń
authors.forename: Jaroslava
authors.initials: J
authors.lastname: Hniliková
authors.forename: Miroslav
authors.initials: M
authors.lastname: Šíša
authors.forename: Dieter
authors.initials: D
authors.lastname: Buyst
authors.forename: Josep
authors.initials: J
authors.lastname: Vilarrasa-Blasi
authors.forename: Anna-Mária
authors.initials: AM
authors.lastname: Szatmári
authors.forename: Daniël
authors.initials: D
authors.lastname: Van Damme
authors.forename: Kiril
authors.initials: K
authors.lastname: Mishev
authors.forename: Mirela-Corina
authors.initials: MC
authors.lastname: Codreanu
authors.forename: Ladislav
authors.initials: L
authors.lastname: Kohout
authors.forename: Miroslav
authors.initials: M
authors.lastname: Strnad
authors.forename: Ana I
authors.initials: AI
authors.lastname: Caño-Delgado
authors.forename: Jiří
authors.initials: J
authors.lastname: Friml
authors.forename: Annemieke
authors.initials: A
authors.lastname: Madder
authors.forename: Eugenia
authors.initials: E
authors.lastname: Russinova
mesh_terms: D017360:Arabidopsis / Q000201:enzymology / Q000378:metabolism* / Q000648:ultrastructure; D029681:Arabidopsis Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D060406:Brassinosteroids / Q000737:chemistry / Q000378:metabolism; D002232:Carbocyanines / Q000737:chemistry*; D002462:Cell Membrane / Q000378:metabolism* / Q000648:ultrastructure; D002777:Cholestanols / Q000737:chemistry / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D004705:Endocytosis; D011992:Endosomes / Q000201:enzymology / Q000378:metabolism / Q000648:ultrastructure; D005456:Fluorescent Dyes / Q000737:chemistry*; D049452:Green Fluorescent Proteins / Q000235:genetics; D007700:Kinetics; D018519:Meristem / Q000201:enzymology / Q000378:metabolism / Q000648:ultrastructure; D018613:Microscopy, Confocal; D015394:Molecular Structure; D010937:Plant Growth Regulators; D011494:Protein Kinases / Q000235:genetics / Q000378:metabolism*; D021381:Protein Transport; D036226:Seedlings / Q000201:enzymology / Q000378:metabolism / Q000648:ultrastructure; D015398:Signal Transduction; D014617:Vacuoles / Q000201:enzymology / Q000378:metabolism / Q000648:ultrastructure
chemical_list: C569686:Alexa Fluor 647; D029681:Arabidopsis Proteins; D060406:Brassinosteroids; D002232:Carbocyanines; D002777:Cholestanols; D005456:Fluorescent Dyes; D010937:Plant Growth Regulators; D049452:Green Fluorescent Proteins; C083545:castasterone; D011494:Protein Kinases; C108270:BRI1 protein, Arabidopsis