| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
|
|
|
|||||||
|
|||||||||
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Membrane Protein Research Group, Departments of
1 Physiology
2 Oncology and Faculty of
3 Pharmacy, University of Alberta, Edmonton, Alberta T6G 2H7, Canada
4 Cross Cancer Institute, Edmonton, Alberta T6G 2H7, Canada
5 School of Biochemistry and Microbiology, University of Leeds, Leeds LS2 9JT, UK
Human concentrative nucleoside transporter 1 (hCNT1) mediates active transport of nucleosides and anticancer and antiviral nucleoside drugs across cell membranes by coupling influx to the movement of Na+ down its electrochemical gradient. The two-microelectrode voltage-clamp technique was used to measure steady-state and presteady-state currents of recombinant hCNT1 produced in Xenopus oocytes. Transport was electrogenic, phloridzin sensitive and specific for pyrimidine nucleosides and adenosine. Nucleoside analogues that induced inwardly directed Na+ currents included the anticancer drugs 5-fluorouridine, 5-fluoro-2'-deoxyuridine, cladribine and cytarabine, the antiviral drugs zidovudine and zalcitabine, and the novel thymidine mimics 1-(2-deoxy-ß-D-ribofuranosyl)-2,4-difluoro-5-methylbenzene and 1-(2-deoxy-ß-D-ribofuranosyl)-2,4-difluoro-5-iodobenzene. Apparent Km values for 5-fluorouridine, 5-fluoro-2'-deoxyuridine and zidovudine were 18, 15 and 450 µM, respectively. hCNT1 was Na+ specific, and the kinetics of steady-state uridine-evoked Na+ currents were consistent with an ordered simultaneous transport model in which Na+ binds first followed by uridine. Membrane potential influenced both ion binding and carrier translocation. The Na+–nucleoside coupling stoichiometry, determined directly by comparing the uridine-induced inward charge movement to [14C]uridine uptake was 1: 1. hCNT1 presteady-state currents were used to determine the fraction of the membrane field sensed by Na+ (61%), the valency of the movable charge (–0.81) and the average number of transporters present in the oocyte plasma membrane (6.8 x 1010 per cell). The hCNT1 turnover rate at –50 mV was 9.6 molecules of uridine transported per second.
(Received 19 May 2004;
accepted after revision 4 June 2004;
first published online 11 June 2004)
Corresponding author J. D. Young: 7–55 Medical Sciences Building University of Alberta, Edmonton, Alberta T6G 2H7, Canada. Email: james.young{at}ualberta.ca
This article has been cited by other articles:
|
|
 |
|
  M. D. Slugoski, K. M. Smith, R. Mulinta, A. M. L. Ng, S. Y. M. Yao, E. L. Morrison, Q. O. T. Lee, J. Zhang, E. Karpinski, C. E. Cass, et al. A Conformationally Mobile Cysteine Residue (Cys-561) Modulates Na+ and H+ Activation of Human CNT3 J. Biol. Chem., September 5, 2008; 283(36): 24922 - 24934. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. D. Slugoski, A. M. L. Ng, S. Y. M. Yao, K. M. Smith, C. C. Lin, J. Zhang, E. Karpinski, C. E. Cass, S. A. Baldwin, and J. D. Young A Proton-mediated Conformational Shift Identifies a Mobile Pore-lining Cysteine Residue (Cys-561) in Human Concentrative Nucleoside Transporter 3 J. Biol. Chem., March 28, 2008; 283(13): 8496 - 8507. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. Y. M. Yao, A. M. L. Ng, M. D. Slugoski, K. M. Smith, R. Mulinta, E. Karpinski, C. E. Cass, S. A. Baldwin, and J. D. Young Conserved Glutamate Residues Are Critically Involved in Na+/Nucleoside Cotransport by Human Concentrative Nucleoside Transporter 1 (hCNT1) J. Biol. Chem., October 19, 2007; 282(42): 30607 - 30617. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 S. Ravera, L. V. Virkki, H. Murer, and I. C. Forster Deciphering PiT transport kinetics and substrate specificity using electrophysiology and flux measurements Am J Physiol Cell Physiol, August 1, 2007; 293(2): C606 - C620. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 E. Errasti-Murugarren, M. Pastor-Anglada, and F. J. Casado Role of CNT3 in the transepithelial flux of nucleosides and nucleoside-derived drugs J. Physiol., August 1, 2007; 582(3): 1249 - 1260. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 D. D.F. Loo, B. A. Hirayama, M. H. Karakossian, A.-K. Meinild, and E. M. Wright Conformational Dynamics of hSGLT1 during Na+/Glucose Cotransport J. Gen. Physiol., December 1, 2006; 128(6): 701 - 720. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
I. M. Larrayoz, A. Fernandez-Nistal, A. Garces, E. Gorraitz, and M. P. Lostao Characterization of the rat Na+/nucleoside cotransporter 2 and transport of nucleoside-derived drugs using electrophysiological methods Am J Physiol Cell Physiol, December 1, 2006; 291(6): C1395 - C1404. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. Zhang, K. M. Smith, T. Tackaberry, X. Sun, P. Carpenter, M. D. Slugoski, M. J. Robins, L. P. C. Nielsen, I. Nowak, S. A. Baldwin, et al. Characterization of the Transport Mechanism and Permeant Binding Profile of the Uridine Permease Fui1p of Saccharomyces cerevisiae J. Biol. Chem., September 22, 2006; 281(38): 28210 - 28221. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 I. Aymerich, F. Foufelle, P. Ferre, F. J. Casado, and M. Pastor-Anglada Extracellular adenosine activates AMP-dependent protein kinase (AMPK) J. Cell Sci., April 15, 2006; 119(8): 1612 - 1621. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J. Zhang, K. M. Smith, T. Tackaberry, F. Visser, M. J. Robins, L. P. C. Nielsen, I. Nowak, E. Karpinski, S. A. Baldwin, J. D. Young, et al. Uridine Binding and Transportability Determinants of Human Concentrative Nucleoside Transporters Mol. Pharmacol., September 1, 2005; 68(3): 830 - 839. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
K. M. Smith, M. D. Slugoski, S. K. Loewen, A. M. L. Ng, S. Y. M. Yao, X.-Z. Chen, E. Karpinski, C. E. Cass, S. A. Baldwin, and J. D. Young The Broadly Selective Human Na+/Nucleoside Cotransporter (hCNT3) Exhibits Novel Cation-coupled Nucleoside Transport Characteristics J. Biol. Chem., July 8, 2005; 280(27): 25436 - 25449. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 L. V. Virkki, I. C. Forster, J. Biber, and H. Murer Substrate interactions in the human type IIa sodium-phosphate cotransporter (NaPi-IIa) Am J Physiol Renal Physiol, May 1, 2005; 288(5): F969 - F981. [Abstract] [Full Text] [PDF]
|
|
| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
