title: A dileucine motif is involved in plasma membrane expression and endocytosis of rat sodium taurocholate cotransporting polypeptide (Ntcp).
abstract: The sodium taurocholate cotransporting polypeptide (Ntcp) is the major uptake transporter for bile salts into liver parenchymal cells, and PKC-mediated endocytosis was shown to regulate the number of Ntcp molecules at the plasma membrane. In this study, mechanisms of Ntcp internalization were analyzed by flow cytometry, immunofluorescence, and Western blot analyses in HepG2 cells. PKC activation induced endocytosis of Ntcp from the plasma membrane by ~30%. Endocytosis of Ntcp was clathrin dependent and was followed by lysosomal degradation. A dileucine motif located in the third intracellular loop of Ntcp was essential for endocytosis but also for processing and plasma membrane targeting, suggesting a dual function of this motif for intracellular trafficking of Ntcp. Mutation of two of five potential phosphorylation sites surrounding the dileucine motif (Thr225 and Ser226) inhibited PKC-mediated endocytosis. In conclusion, we could identify a motif, which is critical for Ntcp plasma membrane localization. Endocytic retrieval protects hepatocytes from elevated bile salt concentrations and is of special interest, because NTCP has been identified as a receptor for the hepatitis B and D virus.
authors.affiliation: Dept. of Gastroenterology, Hepatology and Infectious Diseases, Medical Faculty, Univ. Clinic of the Heinrich-Heine Univ. of Düsseldorf, Moorenstrasse 5, D-40225 Düsseldorf, Germany. kubitz@med.uni-duesseldorf.de.
authors.forename: Claudia
authors.initials: C
authors.lastname: Stross
authors.forename: Stefanie
authors.initials: S
authors.lastname: Kluge
authors.forename: Katrin
authors.initials: K
authors.lastname: Weissenberger
authors.forename: Elisabeth
authors.initials: E
authors.lastname: Winands
authors.forename: Dieter
authors.initials: D
authors.lastname: Häussinger
authors.forename: Ralf
authors.initials: R
authors.lastname: Kubitz
mesh_terms: D020816:Amino Acid Motifs; D000818:Animals; D002462:Cell Membrane / Q000235:genetics / Q000378:metabolism*; D002966:Clathrin / Q000378:metabolism; D004705:Endocytosis / Q000502:physiology*; D005786:Gene Expression Regulation / Q000502:physiology; D056945:Hep G2 Cells; D006801:Humans; D007930:Leucine; D029363:Organic Anion Transporters, Sodium-Dependent / Q000235:genetics / Q000378:metabolism*; D021381:Protein Transport / Q000502:physiology*; D051381:Rats; D012694:Serine; D027981:Symporters / Q000235:genetics / Q000378:metabolism*; D013912:Threonine
chemical_list: D002966:Clathrin; D029363:Organic Anion Transporters, Sodium-Dependent; D027981:Symporters; C071908:sodium-bile acid cotransporter; D013912:Threonine; D012694:Serine; D007930:Leucine
keywords: cholestasis; endocytosis; internalization motif; protein kinase; transporter