Panel A shows representative current traces with a scale of 250 picoamperes and 100 milliseconds. Panel B shows a line graph of normalized current versus voltage comparing the 8C 8A(IL1 25) control and E6A mutant. The x-axis represents voltage in millivolts, and the y-axis represents current divided by maximum current. The half maximal voltage values are 114.5 millivolts for the control and 57.4 millivolts for E6A, with P less than 0.0001 and n equal to 6. Panel C shows representative current traces with the same scale of 250 picoamperes and 100 milliseconds. Panel D shows a line graph of normalized current versus voltage comparing the 8C 8A(IL1 25) control and L136K mutant. The half maximal voltage values are 114.5 millivolts for the control and 40.2 millivolts for L136K, with P less than 0.0001 and n equal to 5. Panel E shows representative current traces recorded at pH 7.4 and pH 6.0, with a scale of 250 picoamperes and 100 milliseconds. Panel F shows normalized current versus voltage at pH 7.4 and pH 6.0. The half maximal voltage values are 68.2 millivolts at pH 7.4 and 57.7 millivolts at pH 6.0, with n equal to 6 for both conditions and P less than 0.0001. Panel G shows percentage inhibition versus DCPIB concentration at pH 7.4 and pH 6.0. The x-axis represents DCPIB concentration in micromolar on a logarithmic scale, and the y-axis represents percentage inhibition. The half maximal inhibitory concentrations are 2.3 micromolar at pH 7.4 and 1.2 micromolar at pH 6.0, with P less than 0.005.
Effects of E6A and L136K mutations and extracellular low pH on tail currents. (A) Representative tail currents of 8C-8A(IL125)-E6A mutant recorded from a cell dialyzed with normal pipette solution and swollen with hypotonic buffer. (B) Mean ± SEM VDI of the tail currents in 8C-8A(IL125)-E6A (n = 5). (C) Representative tail currents of 8C-8A(IL125)-L136K. (D) Mean ± SEM VDI of the tail currents in 8C-8A(IL125)-L136K (n = 5). WT data (black line) from Fig. 3 H is shown in B and D for comparison. (E) Representative tail currents of native VRAC recorded from a WT HCT116 cell dialyzed with normal pipette solution and swollen with hypotonic buffer under normal (pH 7.4) and low pH (pH 6.0) bath conditions. (F) Mean ± SEM pH-induced enhancement of tail current inactivation (n = 6). After full activation and recording of the tail current at pH 7.4, the extracellular buffer was switched to a buffer at pH 6.0. (G) Mean ± SEM DCPIB CRC data recorded at +100 mV from HCT116 cells at pH 7.4 (open symbols, n = 5) and pH 6.0 (blue symbols, n = 6).