title: Clathrin-coated pit-mediated receptor internalization. Role of internalization signals and receptor mobility.
abstract: Most signals controlling receptor-mediated endocytosis have been identified by alteration of sequences present in receptors normally internalized via clathrin-coated pits. In the present work we have reconsidered the factors that control internalization the other way around: i.e. by introducing potential internalization sequences in complement receptor 1 (CR1) which does not preferentially associate with clathrin-coated pits. The analysis of the internalization efficiency of NPxY related motifs generated by substituting His2010 and/or Glu2015 by either Phe or Tyr indicates that FxNPxY is the stronger promoter of endocytosis and that the signal efficiency depends on the presence of aromatic residues (including a tyrosine) at both ends of the -xNPx- motif. Moreover, CR1-tyr (substitution of Glu2015 for Tyr) internalization was superposable to that of a receptor composed of the extracellular and transmembrane domains of CR1 fused to the intracytoplasmic tail of the low density lipoprotein (LDL) receptor (including the FxNPxY motif) (CR1-LDL). When analyzed by fluorescence recovery after photobleaching, the surface mobility of CR1-LDL was decreased as compared with that of either CR1-tyr or CR1-wt, despite a similar association with clathrin-coated pits. The role of receptor mobility in internalization was confirmed by the observation that CR1-tl, with a deletion of the cytoplasmic tail, was more mobile and more efficiently internalized than CR1-wt.
authors.affiliation: Department of Morphology, University Medical Center, University of Geneva, Switzerland.
authors.forename: J P
authors.initials: JP
authors.lastname: Paccaud
authors.forename: W
authors.initials: W
authors.lastname: Reith
authors.forename: B
authors.initials: B
authors.lastname: Johansson
authors.forename: K E
authors.initials: KE
authors.lastname: Magnusson
authors.forename: B
authors.initials: B
authors.lastname: Mach
authors.forename: J L
authors.initials: JL
authors.lastname: Carpentier
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D003034:Coated Pits, Cell-Membrane / Q000502:physiology*; D017931:DNA Primers / Q000737:chemistry; D004705:Endocytosis; D006801:Humans; D066298:In Vitro Techniques; D008560:Membrane Fluidity; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D017463:Receptors, Complement 3b / Q000378:metabolism*; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D013329:Structure-Activity Relationship
chemical_list: D017931:DNA Primers; D017463:Receptors, Complement 3b