Mouse Ae1 E699Q mediates SO42−i/anionoexchange with [SO42−]i-dependent reversal of wild-type pHosensitivity

Abstract

The SLC4A1/AE1 gene encodes the electroneutral Cl−/HCO3−exchanger of erythrocytes and renal type A intercalated cells. AE1 mutations cause familial spherocytic and stomatocytic anemias, ovalocytosis, and distal renal tubular acidosis. The mutant mouse Ae1 polypeptide E699Q expressed in Xenopus oocytes cannot mediate Cl−/HCO3−exchange or36Cl−efflux but exhibits enhanced dual sulfate efflux mechanisms: electroneutral exchange of intracellular sulfate for extracellular sulfate (SO42−i/SO42−oexchange), and electrogenic exchange of intracellular sulfate for extracellular chloride (SO42−i/Cl−oexchange). Whereas wild-type AE1 mediates 1:1 H+/SO42−cotransport in exchange for either Cl−or for the H+/SO42−ion pair, mutant Ae1 E699Q transports sulfate without cotransport of protons, similar to human erythrocyte AE1 in which the corresponding E681 carboxylate has been chemically converted to the alcohol (hAE1 E681OH). We now show that in contrast to the normal cis-stimulation by protons of wild-type AE1-mediated SO42−transport, both SO42−i/Cl−oexchange and SO42−i/SO42−oexchange mediated by mutant Ae1 E699Q are inhibited by acidic pHoand activated by alkaline pHo. hAE1 E681OH displays a similarly altered pHodependence of SO42−i/Cl−oexchange. Elevated [SO42−]iincreases the K1/2of Ae1 E699Q for both extracellular Cl−and SO42−, while reducing inhibition of both exchange mechanisms by acid pHo. The E699Q mutation also leads to increased potency of self-inhibition by extracellular SO42−. Study of the Ae1 E699Q mutation has revealed the existence of a novel pH-regulatory site of the Ae1 polypeptide and should continue to provide valuable paths toward understanding substrate selectivity and self-inhibition in SLC4 anion transporters.