title: Contribution of a tyrosine-based motif to cellular trafficking of wild-type and truncated NPY Y(1) receptors.
abstract: The human NPY Y(1) receptor undergoes fast agonist-induced internalization via clathrin-coated pits then recycles back to the cell membrane. In an attempt to identify the molecular determinants involved in this process, we studied several C-terminal truncation mutants tagged with EFGP. In the absence of agonist, Y(1) receptors lacking the last 32 C-terminal amino acids (Y(1)Δ32) are constitutively internalized, unlike full-length Y(1) receptors. At steady state, internalized Y(1)Δ32 receptors co-localize with transferrin, a marker of early and recycling endosomes. Inhibition of constitutive internalization of Y(1)Δ32 receptors by hypertonic sucrose or by co-expression of Rab5aS34N, a dominant negative form of the small GTPase Rab5a or depletion of all three isoforms of Rab5 indicates the involvement of clathrin-coated pits. In contrast, a truncated receptor lacking the last 42 C-terminal amino acids (Y(1)Δ42) does not constitutively internalize, consistent with the possibility that there is a molecular determinant responsible for constitutive internalization located in the last 10 amino acids of Y(1)Δ32 receptors. We show that the agonist-independent internalization of Y(1)Δ32 receptors involves a tyrosine-based motif YXXΦ. The potential role of this motif in the behaviour of full-length Y(1) receptors has also been explored. Our results indicate that a C-terminal tyrosine-based motif is critical for the constitutive internalization of truncated Y(1)Δ32 receptors. We suggest that this motif is masked in full-length Y(1) receptors which do not constitutively internalize in the absence of agonist.
authors.affiliation: Institut de Recherche de l'Ecole de Biotechnologie de Strasbourg, FRE 3211, Ecole Supérieure de Biotechnologie de Strasbourg, Boulevard Sébastien Brant, Illkirch, France.
authors.forename: Sandra
authors.initials: S
authors.lastname: Lecat
authors.forename: Moussa
authors.initials: M
authors.lastname: Ouédraogo
authors.forename: Thomas
authors.initials: T
authors.lastname: Cherrier
authors.forename: Fanny
authors.initials: F
authors.lastname: Noulet
authors.forename: Philippe
authors.initials: P
authors.lastname: Rondé
authors.forename: Nicole
authors.initials: N
authors.lastname: Glasser
authors.forename: Jean-Luc
authors.initials: JL
authors.lastname: Galzi
authors.forename: Yves
authors.initials: Y
authors.lastname: Mely
authors.forename: Kenneth
authors.initials: K
authors.lastname: Takeda
authors.forename: Bernard
authors.initials: B
authors.lastname: Bucher
mesh_terms: D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D002966:Clathrin / Q000737:chemistry / Q000378:metabolism; D057809:HEK293 Cells; D006801:Humans; D007700:Kinetics; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009478:Neuropeptide Y / Q000494:pharmacology; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000378:metabolism; D017476:Receptors, Neuropeptide Y / Q000819:agonists / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction; D014168:Transferrin / Q000378:metabolism; D014443:Tyrosine / Q000378:metabolism*; D020696:rab5 GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism
chemical_list: D002966:Clathrin; D009478:Neuropeptide Y; D034741:RNA, Small Interfering; D017476:Receptors, Neuropeptide Y; D014168:Transferrin; C097212:neuropeptide Y-Y1 receptor; D014443:Tyrosine; D020696:rab5 GTP-Binding Proteins