Figure S1.

Peak KCNQ1 currents after ML277 addition and response to HMR. (A) Diary plots of peak KCNQ1 current over time after 1 µM ML277 addition, normalized to last control level for eight different cells. Cells were held at −80 mV and pulsed to +60 mV for 2 s, then to −40 mV for 1 s repeatedly with an interpulse interval of 15 s. In a few examples (red triangles, yellow circles), after ML277 has its effect, the current begins to decrease again before stabilizing, and while the decrease did not always happen in the recording time frame, it suggests that ML277 might not act as a PIP2 mimetic given that PIP2 has previously been shown to prevent rundown of whole-cell currents (Li et al., 2011; Loussouarn et al., 2003). (B) Concentration-response curves for HMR inhibition of KCNQ1 currents under different experimental conditions; KCNQ1 pulsed to +60 mV and HMR added after washout of ML277 (red circles, IC₅₀ = 3.44 µM); KCNQ1 pulsed to +60 mV and HMR added with ML277 remaining in solution (magenta triangles, IC₅₀ = 5.88 µM); KCNQ1 pulsed to −20 mV and HMR added with ML277 remaining in solution (green triangles, IC₅₀ = 4.86 µM); and KCNQ1 in control bath solution pulsed to +60 mV (blue circles, IC₅₀ = 2.54 µM), n = 2–6. (C) Paired peak tail current amplitudes before and after exposure to 1 µM ML277. Protocol was as in A. (D) Chart of peak KCNQ1 tail current amplitudes in ML277 divided by peak current in control conditions with four different test protocols as labeled. (E) Diary plots of KCNQ1 peak currents showing response to increasing concentrations of HMR under the different experimental conditions detailed in B. A pause in the diary plot is shown where a voltage-clamp protocol was run before beginning the 10-min washout. (F) Top: G-V plot of KCNQ1 before (blue circles) and after exposure to 0.1% DMSO (cyan circles). V_1/2 of activation and slope values was −22.5 ± 1.5 mV and 12.8 ± 2.0 for control, −23.7 ± 1.8 mV and 14.5 ± 1.8 in DMSO, n = 4 for each. Cells were held at −90 mV and pulsed from −90 to +80 for 4 s, then to −40 mV for 0.9 s, with an interpulse interval of 15 s. Bottom: Plot of peak tail currents in 0.1% DMSO divided by control values after a 4-s, +60 mV pulse. (G) Ensemble averages of 53 sweeps of KCNQ1 treated with 1 µM ML277 (red) and 53 sweeps of the same cell after treatment with 20 µM HMR. Protocol is shown above. The interpulse interval was 10 s. All error bars in the figure denote mean ± SEM.