title: Sorting Motifs Involved in the Trafficking and Localization of the PIN1 Auxin Efflux Carrier.
abstract: In contrast with the wealth of recent reports about the function of μ-adaptins and clathrin adaptor protein (AP) complexes, there is very little information about the motifs that determine the sorting of membrane proteins within clathrin-coated vesicles in plants. Here, we investigated putative sorting signals in the large cytosolic loop of the Arabidopsis (Arabidopsis thaliana) PIN-FORMED1 (PIN1) auxin transporter, which are involved in binding μ-adaptins and thus in PIN1 trafficking and localization. We found that Phe-165 and Tyr-280, Tyr-328, and Tyr-394 are involved in the binding of different μ-adaptins in vitro. However, only Phe-165, which binds μA(μ2)- and μD(μ3)-adaptin, was found to be essential for PIN1 trafficking and localization in vivo. The PIN1:GFP-F165A mutant showed reduced endocytosis but also localized to intracellular structures containing several layers of membranes and endoplasmic reticulum (ER) markers, suggesting that they correspond to ER or ER-derived membranes. While PIN1:GFP localized normally in a μA (μ2)-adaptin mutant, it accumulated in big intracellular structures containing LysoTracker in a μD (μ3)-adaptin mutant, consistent with previous results obtained with mutants of other subunits of the AP-3 complex. Our data suggest that Phe-165, through the binding of μA (μ2)- and μD (μ3)-adaptin, is important for PIN1 endocytosis and for PIN1 trafficking along the secretory pathway, respectively.
authors.affiliation: Departamento de Bioquímica y Biología Molecular, Facultad de Farmacia, Universitat de Valencia, 46100 Burjassot, Spain (G.S.-A., E.S.-O., J.M.B.-O., F.A., M.J.M.);Institute of Science and Technology Austria, 3400 Klostenburg, Austria (M.N., A.O.M., R.T., J.F.); andSchool of Life Sciences, Centre for Cell and Developmental Biology and State Key Laboratory of Agrobiotechnology, Chinese University of Hong Kong, Hong Kong, China (C.G., L.J.).
authors.forename: Gloria
authors.initials: G
authors.lastname: Sancho-Andrés
authors.affiliation: Departamento de Bioquímica y Biología Molecular, Facultad de Farmacia, Universitat de Valencia, 46100 Burjassot, Spain (G.S.-A., E.S.-O., J.M.B.-O., F.A., M.J.M.);Institute of Science and Technology Austria, 3400 Klostenburg, Austria (M.N., A.O.M., R.T., J.F.); andSchool of Life Sciences, Centre for Cell and Developmental Biology and State Key Laboratory of Agrobiotechnology, Chinese University of Hong Kong, Hong Kong, China (C.G., L.J.).
authors.forename: Esther
authors.identifier: 0000-0002-2352-606X
authors.initials: E
authors.lastname: Soriano-Ortega
authors.affiliation: Departamento de Bioquímica y Biología Molecular, Facultad de Farmacia, Universitat de Valencia, 46100 Burjassot, Spain (G.S.-A., E.S.-O., J.M.B.-O., F.A., M.J.M.);Institute of Science and Technology Austria, 3400 Klostenburg, Austria (M.N., A.O.M., R.T., J.F.); andSchool of Life Sciences, Centre for Cell and Developmental Biology and State Key Laboratory of Agrobiotechnology, Chinese University of Hong Kong, Hong Kong, China (C.G., L.J.).
authors.forename: Caiji
authors.initials: C
authors.lastname: Gao
authors.affiliation: Departamento de Bioquímica y Biología Molecular, Facultad de Farmacia, Universitat de Valencia, 46100 Burjassot, Spain (G.S.-A., E.S.-O., J.M.B.-O., F.A., M.J.M.);Institute of Science and Technology Austria, 3400 Klostenburg, Austria (M.N., A.O.M., R.T., J.F.); andSchool of Life Sciences, Centre for Cell and Developmental Biology and State Key Laboratory of Agrobiotechnology, Chinese University of Hong Kong, Hong Kong, China (C.G., L.J.).
authors.forename: Joan Miquel
authors.initials: JM
authors.lastname: Bernabé-Orts
authors.affiliation: Departamento de Bioquímica y Biología Molecular, Facultad de Farmacia, Universitat de Valencia, 46100 Burjassot, Spain (G.S.-A., E.S.-O., J.M.B.-O., F.A., M.J.M.);Institute of Science and Technology Austria, 3400 Klostenburg, Austria (M.N., A.O.M., R.T., J.F.); andSchool of Life Sciences, Centre for Cell and Developmental Biology and State Key Laboratory of Agrobiotechnology, Chinese University of Hong Kong, Hong Kong, China (C.G., L.J.).
authors.forename: Madhumitha
authors.identifier: 0000-0002-8600-0671
authors.initials: M
authors.lastname: Narasimhan
authors.affiliation: Departamento de Bioquímica y Biología Molecular, Facultad de Farmacia, Universitat de Valencia, 46100 Burjassot, Spain (G.S.-A., E.S.-O., J.M.B.-O., F.A., M.J.M.);Institute of Science and Technology Austria, 3400 Klostenburg, Austria (M.N., A.O.M., R.T., J.F.); andSchool of Life Sciences, Centre for Cell and Developmental Biology and State Key Laboratory of Agrobiotechnology, Chinese University of Hong Kong, Hong Kong, China (C.G., L.J.).
authors.forename: Anna Ophelia
authors.initials: AO
authors.lastname: Müller
authors.affiliation: Departamento de Bioquímica y Biología Molecular, Facultad de Farmacia, Universitat de Valencia, 46100 Burjassot, Spain (G.S.-A., E.S.-O., J.M.B.-O., F.A., M.J.M.);Institute of Science and Technology Austria, 3400 Klostenburg, Austria (M.N., A.O.M., R.T., J.F.); andSchool of Life Sciences, Centre for Cell and Developmental Biology and State Key Laboratory of Agrobiotechnology, Chinese University of Hong Kong, Hong Kong, China (C.G., L.J.).
authors.forename: Ricardo
authors.identifier: 0000-0003-1169-731X
authors.initials: R
authors.lastname: Tejos
authors.affiliation: Departamento de Bioquímica y Biología Molecular, Facultad de Farmacia, Universitat de Valencia, 46100 Burjassot, Spain (G.S.-A., E.S.-O., J.M.B.-O., F.A., M.J.M.);Institute of Science and Technology Austria, 3400 Klostenburg, Austria (M.N., A.O.M., R.T., J.F.); andSchool of Life Sciences, Centre for Cell and Developmental Biology and State Key Laboratory of Agrobiotechnology, Chinese University of Hong Kong, Hong Kong, China (C.G., L.J.).
authors.forename: Liwen
authors.initials: L
authors.lastname: Jiang
authors.affiliation: Departamento de Bioquímica y Biología Molecular, Facultad de Farmacia, Universitat de Valencia, 46100 Burjassot, Spain (G.S.-A., E.S.-O., J.M.B.-O., F.A., M.J.M.);Institute of Science and Technology Austria, 3400 Klostenburg, Austria (M.N., A.O.M., R.T., J.F.); andSchool of Life Sciences, Centre for Cell and Developmental Biology and State Key Laboratory of Agrobiotechnology, Chinese University of Hong Kong, Hong Kong, China (C.G., L.J.).
authors.forename: Jiří
authors.identifier: 0000-0002-8302-7596
authors.initials: J
authors.lastname: Friml
authors.affiliation: Departamento de Bioquímica y Biología Molecular, Facultad de Farmacia, Universitat de Valencia, 46100 Burjassot, Spain (G.S.-A., E.S.-O., J.M.B.-O., F.A., M.J.M.);Institute of Science and Technology Austria, 3400 Klostenburg, Austria (M.N., A.O.M., R.T., J.F.); andSchool of Life Sciences, Centre for Cell and Developmental Biology and State Key Laboratory of Agrobiotechnology, Chinese University of Hong Kong, Hong Kong, China (C.G., L.J.) fernando.aniento@uv.es mariajesus.marcote@uv.es.
authors.forename: Fernando
authors.identifier: 0000-0003-3553-585X
authors.initials: F
authors.lastname: Aniento
authors.affiliation: Departamento de Bioquímica y Biología Molecular, Facultad de Farmacia, Universitat de Valencia, 46100 Burjassot, Spain (G.S.-A., E.S.-O., J.M.B.-O., F.A., M.J.M.);Institute of Science and Technology Austria, 3400 Klostenburg, Austria (M.N., A.O.M., R.T., J.F.); andSchool of Life Sciences, Centre for Cell and Developmental Biology and State Key Laboratory of Agrobiotechnology, Chinese University of Hong Kong, Hong Kong, China (C.G., L.J.) fernando.aniento@uv.es mariajesus.marcote@uv.es.
authors.forename: María Jesús
authors.initials: MJ
authors.lastname: Marcote
mesh_terms: D033983:Adaptor Protein Complex mu Subunits / Q000235:genetics / Q000378:metabolism; D017360:Arabidopsis / Q000235:genetics / Q000378:metabolism*; D029681:Arabidopsis Proteins / Q000235:genetics / Q000378:metabolism*; D002966:Clathrin / Q000378:metabolism; D003600:Cytosol / Q000378:metabolism; D004705:Endocytosis / Q000235:genetics; D004721:Endoplasmic Reticulum / Q000235:genetics / Q000378:metabolism; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D020662:Guanine Nucleotide Exchange Factors / Q000235:genetics; D026901:Membrane Transport Proteins / Q000235:genetics / Q000378:metabolism*; D009154:Mutation; D010649:Phenylalanine / Q000235:genetics; D030821:Plants, Genetically Modified; D021382:Protein Sorting Signals / Q000235:genetics; D021381:Protein Transport
chemical_list: D033983:Adaptor Protein Complex mu Subunits; D029681:Arabidopsis Proteins; D002966:Clathrin; C522273:GNL1 protein, Arabidopsis; D020662:Guanine Nucleotide Exchange Factors; D026901:Membrane Transport Proteins; C116363:PIN1 protein, Arabidopsis; D021382:Protein Sorting Signals; D049452:Green Fluorescent Proteins; D010649:Phenylalanine