High affinity sodium-dependent nucleobase transport in cultured renal epithelial cells (LLC-PK1).
Two distinct transporters for nucleobases have been characterized in LLC-PK1 cells. The first system accumulates hypoxanthine against a concentration gradient in the presence of sodium. The sodium-dependent uptake of hypoxanthine was saturable at 22 degrees C with a Km value of 0.79 +/- 0.43 microM, a Vmax of 15 +/- 4 pmol/mg protein/60 s, and a Na+:hypoxanthine coupling stoichiometry of 1.27 +/- 0.20. Uptake of hypoxanthine was inhibited by 5-fluorouracil, uracil, thymine, and guanine (Ki values 3-6 microM). Adenine and nucleosides were without effect. Using cell monolayers grown on a permeable filter support, Na(+)-dependent hypoxanthine uptake occurred preferentially from the apical surface. The second system exhibited no cation specificity and was saturable with a low affinity for hypoxanthine (Km of 124 +/- 22 microM) and a high Vmax of 275 +/- 38 pmol/mg protein/60 s. Adenine and guanine inhibited Na(+)-independent hypoxanthine uptake (Ki values 30 +/- 15 and 18 +/- 7 microM, respectively). Other nucleobases and nucleosides exhibited little or no inhibition of equilibrative hypoxanthine influx. Dipyridamole, dilazep, and phloridzin were effective inhibitors of Na(+)-dependent hypoxanthine uptake but had little effect on the Na(+)-independent flux. This study represents the first direct demonstration of a unique high affinity Na+ nucleobase co-transporter system in cultured animal cells.
Research School of Biosciences, University of Kent, Canterbury, United Kingdom.
D A
DA
Griffith
S M
SM
Jarvis
D000818:Animals; D001692:Biological Transport / Q000187:drug effects; D002414:Cations, Monovalent / Q000494:pharmacology; D002460:Cell Line; D004109:Dilazep / Q000494:pharmacology; D004176:Dipyridamole / Q000494:pharmacology; D004848:Epithelium / Q000378:metabolism; D019271:Hypoxanthine; D007042:Hypoxanthines / Q000378:metabolism*; D007668:Kidney; D007700:Kinetics; D009705:Nucleosides / Q000494:pharmacology; D011687:Purines / Q000494:pharmacology; D011743:Pyrimidines / Q000494:pharmacology; D012964:Sodium / Q000494:pharmacology*; D013868:Thioinosine / Q000031:analogs & derivatives / Q000494:pharmacology; D014316:Tritium
D002414:Cations, Monovalent; D007042:Hypoxanthines; D009705:Nucleosides; D011687:Purines; D011743:Pyrimidines; D014316:Tritium; D019271:Hypoxanthine; D013868:Thioinosine; D004176:Dipyridamole; D012964:Sodium; D004109:Dilazep; C001789:4-nitrobenzylthioinosine