Fig. 6.

MATE1 mediates electroneutral H⁺/TEA exchange. (A) Presence of wild-type and E273Q mutated hMATE1 in HEK293 cells, as revealed by indirect immunofluorescence microscopy. No immunoreactivity was observed in a mock control, which was from HEK293 cells transfected with pcDNA3.1 vector alone. (B) Time course of TEA (50 μM) uptake at pH 8.0 by HEK293 cells expressing hMATE1, the E273Q mutant, or a mock control. (C) Dose dependence of TEA uptake at pH 8.0. The values obtained at the indicated concentrations from the mock control cells were subtracted from the corresponding values obtained from cells expressing wild-type hMATE1. (D) pH dependence of TEA uptake. HEK293 cells expressing wild-type hMATE or a mock control were incubated at the indicated pH, and TEA uptake was then measured. (E) The effect of Na⁺ on TEA uptake was examined in buffer containing 65 mM KCl and 65 mM NaCl (control) or in buffer containing 130 mM KCl (Na⁺ free). The requirement of a membrane potential or pH gradient for TEA uptake was also examined at pH 8.0 in the absence or presence of 10 mM ammonium chloride, 5 μM nigericin, 10 μM SF6847, or 5 μM valinomycin in buffer containing 65 mM KCl and 65 mM NaCl (control). (F) pH-dependent extrusion of TEA. hMATE1-expressing cells were incubated with 50 μM radiolabeled TEA as in Fig. 2B for 10 min. Then, the cells were transferred to buffer with the indicated pH (time 0) and incubated for a further 10 min and the remaining radioactivity then assayed. Error bars are the standard deviation of three samples.