title: Sorting signals in the cytosolic tail of membrane proteins involved in the interaction with plant ARF1 and coatomer.
abstract: In mammals and yeast, a cytosolic dilysine motif is critical for endoplasmic reticulum (ER) localization of type I membrane proteins. Retrograde transport of type I membrane proteins containing dilysine motifs at their cytoplasmic carboxy (C)-terminal tail involves the interaction of these motifs with the COPI coat. The C-terminal dilysine motif has also been shown to confer ER localization to type I membrane proteins in plant cells. Using in vitro binding assays, we have analyzed sorting motifs in the cytosolic tail of membrane proteins, which may be involved in the interaction with components of the COPI coat in plant cells. We show that a dilysine motif in the -3,-4 position (relative to the cytosolic C-terminus) recruits in a very specific manner all the subunits of the plant coatomer complex. Lysines cannot be replaced by arginines or histidines to bind plant coatomer. A diphenylalanine motif in the -7,-8 position, which by itself has a low ability to bind plant coatomer, shows a clear cooperativity with the dilysine motif. Both dilysine and diphenylalanine motifs are present in the cytosolic tail of several proteins of the p24 family of putative cargo receptors, which has several members in plant cells. The cytosolic tail of a plant p24 protein is shown to recruit not only coatomer but also ADP ribosylation factor 1 (ARF1), a process which depends on both dilysine and diphenylalanine motifs. ARF1 binding increases twofold upon treatment with brefeldin A (BFA) and is completely abolished upon treatment with GTPgammaS, suggesting that ARF1 can only interact with the cytosolic tail of p24 proteins in its GDP-bound form.
authors.affiliation: Departamento de Bioquímica y Biología Molecular, Facultad de Farmacia, Universidad de Valencia, Avenida Vicente Andrés Estellés s/n, E-46100 Burjassot, Valencia, Spain.
authors.forename: Inmaculada
authors.initials: I
authors.lastname: Contreras
authors.forename: Elena
authors.initials: E
authors.lastname: Ortiz-Zapater
authors.forename: Fernando
authors.initials: F
authors.lastname: Aniento
mesh_terms: D020823:ADP-Ribosylation Factor 1 / Q000737:chemistry / Q000378:metabolism*; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D001665:Binding Sites; D020756:Coatomer Protein / Q000737:chemistry / Q000378:metabolism*; D007700:Kinetics; D008565:Membrane Proteins / Q000502:physiology*; D012275:Oryza / Q000502:physiology*; D010446:Peptide Fragments / Q000737:chemistry; D010940:Plant Proteins / Q000737:chemistry / Q000378:metabolism*; D021122:Protein Subunits / Q000737:chemistry / Q000378:metabolism; D011523:Protoplasts / Q000502:physiology; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D015398:Signal Transduction / Q000502:physiology*
chemical_list: D020756:Coatomer Protein; D008565:Membrane Proteins; D010446:Peptide Fragments; D010940:Plant Proteins; D021122:Protein Subunits; D020823:ADP-Ribosylation Factor 1