FIGURE 4.

Cysteine mutations in the LEL differentially affect transport activity of KCC2 and KCC4. HEK-293 cells were assayed for ⁸⁶Rb⁺ uptake 48 h after transfection with the constructs indicated below the columns. An empty vector was used for mock transfection. A, ⁸⁶Rb⁺ uptake of the quadruple cysteine mutant KCC2_q (C287S,C302L,C322S,C331L) resulted in a decrease of the transport activity compared with KCC2_wt (p < 0.001). Treatment with 1 mm NEM did not increase the transport activity of the quadruple mutant in contrast to its effect on KCC2_wt. B, KCC4-transfected cells revealed increased transport activity in relation to mock-transfected cells. ⁸⁶Rb⁺ uptake of KCC4_wt and the quadruple mutant KCC4_q (C308S,C323L,C343S,C352L) is similar. The plot depicts the mean of three experiments ± S.D. (error bars). ***, p < 0.001; n.s., not significant.