Cell-specific basolateral membrane sorting of the human liver Na(+)-dependent bile acid cotransporter.
The human Na(+)-taurocholate cotransporting polypeptide (Ntcp) is located exclusively on the basolateral membrane of hepatocyte, but the mechanisms underlying its membrane sorting domain have not been fully elucidated. In the present study, a green fluorescent protein-fused human NTCP (NTCP-GFP) was constructed using the polymerase chain reaction and was stably transfected into Madin-Darby canine kidney (MDCK) and Caco-2 cells. Taurocholate uptake studies and confocal microscopy demonstrated that the polarity of basolateral surface expression of NTCP-GFP was maintained in MDCK cells but was lost in Caco-2 cells. Nocodazole (33 microM), an agent that causes microtubular depolymerization, partially disrupted the basolateral localization of NTCP-GFP by increasing apical surface expression to 33.5% compared with untreated cells (P < 0.05). Brefeldin A (BFA; 1-2 microM) disrupted the polarized basolateral localization of NTCP, but monensin (1.4 microM) had no affect on NTCP-GFP localization. In addition, low-temperature shift (20 degrees C) did not affect the polarized basolateral surface sorting of NTCP-GFP and repolarization of this protein after BFA interruption. In summary, these data suggest that the polarized basolateral localization of human NTCP is cell specific and is mediated by a novel sorting pathway that is BFA sensitive and monensin and low-temperature shift insensitive. The process may also involve microtubule motors.
Sun
A Q
AQ
Department of Pediatrics, Mount Sinai School of Medicine, One Gustave L. Levy Place, New York, NY 10029, USA.
Swaby
I
I
Xu
S
S
Suchy
F J
FJ
D000818:Animals; D019556:COS Cells; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D004285:Dogs; D049452:Green Fluorescent Proteins; D006801:Humans; D007425:Intracellular Membranes / Q000378:metabolism*; D008099:Liver / Q000166:cytology / Q000378:metabolism*; D008164:Luminescent Proteins / Q000235:genetics; D026901:Membrane Transport Proteins; D029363:Organic Anion Transporters, Sodium-Dependent; D021382:Protein Sorting Signals / Q000502:physiology*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D027981:Symporters
D002352:Carrier Proteins; D008164:Luminescent Proteins; D026901:Membrane Transport Proteins; D029363:Organic Anion Transporters, Sodium-Dependent; D021382:Protein Sorting Signals; D011993:Recombinant Fusion Proteins; D027981:Symporters; C071908:sodium-bile acid cotransporter; D049452:Green Fluorescent Proteins