Nucleobase transport in opossum kidney epithelial cells and Xenopus laevisoocytes: the characterisation, structure-activity relationship of uracil analogues and oocyte expression studies of sodium-dependent and -independenthypoxanthine uptake

The characteristics of hypoxanthine transport were examined in opossum kidn ey (OK) epithelial cells and Xenopus laevis oocytes. In both cell types hyp oxanthine influx was mediated by two distinct transport systems: a high-aff inity Na+-dependent system and a Na+-independent transporter. Na+-dependent hypoxanthine transport in OK cells was saturable (K-m 0.78 +/- 0.29 mu M) and was inhibited by guanine, uracil, thymine and 5-fluorouracil (K-i value s 0.5-7 mu M), whereas adenine had no effect. Substitutions at the 2- and 4 -position had a marked effect on the ability of uracil to inhibit Na+/hypox anthine influx by OK cells revealing that an oxo group at both the 2- and 4 -positions of uracil is required for interacting with the transporter. The properties of Na+-dependent hypoxanthine influx in oocytes were similar to those observed in OK cells. In particular, xanthine and oxypurinol inhibite d hypoxanthine influx, a characteristic not observed previously for the Na/nucleobase carrier in pig LLC-PK1 renal cells. Na+-independent hypoxanthin e influx in OK cells and oocytes was of a lower affinity (K-m 90-180 mu M). Adenine and guanine inhibited Na+-independent hypoxanthine flux in OK cell s, but had no effect in oocytes. Injection of LLC-PK1 mRNA into oocytes res ulted in a 1.5-fold stimulation of Na+/hypoxanthine flux over water-injecte d oocytes. These results reveal further heterogeneity in Na+/nucleobase cot ransporters. (C) 1999 Elsevier Science B.V. All rights reserved.