MOAT4, a novel multispecific organic-anion transporter for glucuronides and mercapturates in mouse L1210 cells and human erythrocytes

Glucuronides and mercapturates were examined as possible high-affinity substrates for a low-affinity ATP-dependent transport system for 2,4-dinitrophenyl S-glutathione (DNP-SG) in mouse L1210 cells. Initial inhibitor studies with inside-out vesicles revealed that the low-affinity transport of [H-3]DNP-SG (K-m 450 mu M) exhibits a high sensitivity to N-acetyl 2,4-dinitrophenyl cysteine (NAc-DNP-Cys) (K-i 5.0 mu M) and alpha-naphthyl beta-D-glucuronide (naphthyl glucuronide) (K-i 8.5 mu M). Direct transport measurements showed the presence of ATP-dependent uptake activities for NAc-DNP-[S-35]Cys and naphthyl [C-14] glucuronide, and K-m values for half-maximal transport were comparable to the K-i values of these compounds for inhibition of [H-3]DNP-SG transport. Transport of [H-3]DNP-SG, NAc-DNP-[S-35]Cys and naphthyl [C-14]glucuronide each showed the same sensitivity to various anions and anion conjugates. Inhibition was competitive and was most potent for bilirubin ditaurate, indoprofen, 4-biphenylacetic acid, 4-acridine 4-beta-D-glucuronide, N-acetyl leukotriene E(4), 17 beta-oestradiol 3 beta-D-glucuronide and taurolithocholate 3-sulphate. Inside-out vesicles from human erythrocytes contain a comparable ATP-dependent transport system. These results show that NAc-DNP-Cys and naphthyl glucuronide are high-affinity substrates for a single system identified previously as a low-affinity transporter of DNP-SG. Substrate and inhibitor studies identify this system as a novel multispecific organic-anion transport system (MOAT4) that accommodates glucuronides and mercapturates and is distinct from other MOAT transporters. Human erythrocytes contain an additional ATP-dependent system for NAc-DNP-Cys (K-m 33 mu M) that does not transport monoglucuronides.