ORGANIC ANION TRANSPORTING POLYPEPTIDE MEDIATES ORGANIC ANION HCO3- EXCHANGE/

Organic anion transporting polypeptide (oatp) is an integral membrane protein cloned from rat liver that mediates Na+-independent transport of organic anions such as sulfobromophthalein and taurocholic acid, Pr evious studies in rat hepatocytes suggested that organic anion uptake is associated with base exchange, To better characterize the mechanism of oatp-mediated organic anion uptake, we examined transport of tauro cholate in a HeLa cell line stably transfected with oatp under the reg ulation of a zinc-inducible promoter (Shi, X., Bai, S., Forci, A. C., Burk, R. D., Jacquemin, E., Hagenbuch, B., Meier, P. J., and Wolkoff, A. W. (1995) J. Biol. Chem. 270, 25591-25595). Whereas noninduced tran sfected cells showed virtually no uptake of [H-3]taurocholate, tauroch olate uptake by induced cells was Na+-independent and saturable (K-m = 19.4 +/- 3.3 mu M; V-max = 62.2 +/- 1.4 pmol/min/mg protein; n = 3). To test whether organic anion transport is coupled to HCO3- extrusion, we compared the rates of taurocholate-dependent HCO3- efflux from alk ali-loaded noninduced and induced cells. Monolayers grown on glass cov erslips were loaded with the pH-sensitive dye 2',7'-bis(carboxyethyl)- 5(6)-carboxyfluorescein; intracellular pH (pH(I)) was measured by exci tation ratio fluorometry, Noninduced and induced cells were alkalinize d to an equivalent pH(i) (similar to 7.7) by transient exposure to a 5 0 mM HCO3- Cl--free solution. In the absence of extracellular Cl- and taurocholate , isohydric reduction of superfusate HCO3- concentration from 50 to 25 mM resulted in an insignificant change in pH(i) over tim e (dpH(i)/dt) in both groups. Addition of 25 mu M taurocholate to the superfusate led to a rapid fall in pH(i) in induced (-0.037 +/- 0.011 pH units/min to pH(i) of 7.41 +/- 0.14) but not in noninduced (0.003 /- 0.006 pH units/min to pH(i) of 7.61 +/- 0.08) cells (p < 0.03). The se data indicate that oatp-mediated taurocholate transport is Na+-inde pendent, saturable, and accompanied by HCO, exchange, We conclude that organic anion/base exchange is an important, potentially regulatable component of oatp function.