Protein kinase C induces endocytosis of the sodium taurocholate cotransporting polypeptide.
Bile salts influence signaling and metabolic pathways. In hepatocytes, the sodium taurocholate cotransporting polypeptide (Ntcp) is a major determinant of intracellular bile salt levels. Short-term downregulation of Ntcp is not well characterized to date. FLAG and enhanced green fluorescent protein (EGFP) tags were cloned to the extra- and intracellular termini of Ntcp. Endocytosis of Ntcp in transfected HepG2 cells was visualized by fluorescence of EGFP, and membrane surface expression of Ntcp was quantified by flow cytometry with fluorochrome-labeled FLAG antibodies. Activation of protein kinase C (PKC) by phorbolester or thymeleatoxin an activator of Ca(2+)-dependent conventional PKCs (cPKCs), induced endocytosis of Ntcp, whereas the Na(+)-K(+)-ATPase remained in the plasma membrane. The PKC inhibitor BIM I and the cPKC-selective inhibitor Gö6976 abolished PMA-induced endocytosis. Because of this internalization, cell surface expression of Ntcp was reduced by 36 +/- 7%, bile salt uptake was decreased by 25%, and taurolithocholate sulfate-induced cell toxicity was prevented. In conclusion, Ca(2+)-dependent PKCs induce vesicular retrieval of Ntcp, thereby reducing bile salt uptake. This mechanism may protect hepatocytes from toxic intracellular bile salt concentrations.
Dept. of Gastroenterology, Hepatology and Infectiology, Heinrich-Heine Univ. of Düsseldorf, Germany.
Claudia
C
Stross
Angelika
A
Helmer
Katrin
K
Weissenberger
Boris
B
Görg
Verena
V
Keitel
Dieter
D
Häussinger
Ralf
R
Kubitz
D000818:Animals; D001647:Bile Acids and Salts / Q000378:metabolism; D019610:Cytoprotection; D004705:Endocytosis / Q000502:physiology*; D004789:Enzyme Activation; D005434:Flow Cytometry; D005456:Fluorescent Dyes; D049452:Green Fluorescent Proteins; D056945:Hep G2 Cells; D022781:Hepatocytes / Q000378:metabolism*; D006801:Humans; D029363:Organic Anion Transporters, Sodium-Dependent / Q000235:genetics / Q000378:metabolism*; D009994:Osmolar Concentration; D011493:Protein Kinase C / Q000378:metabolism*; D051381:Rats; D027981:Symporters / Q000235:genetics / Q000378:metabolism*; D014018:Tissue Distribution; D014162:Transfection
D001647:Bile Acids and Salts; D005456:Fluorescent Dyes; D029363:Organic Anion Transporters, Sodium-Dependent; D027981:Symporters; C485184:enhanced green fluorescent protein; C071908:sodium-bile acid cotransporter; D049452:Green Fluorescent Proteins; D011493:Protein Kinase C