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. 2011 Sep 27;286(46):39760–39767. doi: 10.1074/jbc.M111.289090

FIGURE 1.

FIGURE 1.

PMA and membrane depolarization increase KCNQ2/Q3 currents through PI4 kinase β-mediated synthesis of PIP2. A, PMA and membrane depolarization (0 mV and high K+ solution, ND96-K) augment the amplitude of KCNQ2/Q3 currents expressed in Xenopus oocytes. The KCNQ2/Q3 currents were induced by the protocol shown at the top using the TEVC technique. The two current traces (before and 10 min) represent the current traces recorded before and after 10 min treatments of the oocytes with ND96 solution (control), 0 mV (depolarization), ND96-K solution (depolarization), or PMA (50 nm). During treatments, the oocytes were constantly held at −80 mV. B, time course for the effects of PMA (50 nm), 4α-PMA (50 nm), and solvent DMSO on normalized KCNQ2/Q3 currents. The KCNQ currents were measured at +40 mV. The 2 s depolarization at +40 mV was always followed by a longer (1 min) holding potential of −80 mV to avoid the effect of the depolarization. C, summary data for fold increase of KCNQ2/Q3 current after the 10 min treatments described in A (0 mV, 1.31 ± 0.04, n = 8), (ND96-K, 1.38 ± 0.03, n = 10), (PMA, 1.98 ± 0.06, n = 8). **, p < 0.01 compared with ND96 group. D, TLC method was used to measure PIP2 and PIP levels in oocytes from the control group and from the PMA-treated group (50 nm, 15 min). Triplicate samples from a single experiment are shown in the left panel. Summary data from three independent experiments are shown in the right panel. *, p < 0.05 compared with the control group. E, dsRNA for PI4 kinase β blocked PMA- or ND96-K (depolarization)-induced enhancement of PI4 kinase β (PI4K β) expression. The left panel shows the Western blot results for PI4Kβ expression after treatment with control, PMA, ND96K, dsRNA, PMA plus dsRNA, or dsRNA plus ND96K. The right panel shows the relative intensity of the blots from four independent experiments. *, p < 0.05 compared with control. F, in the oocytes injected with dsRNA for PI4K β, PMA increased KCNQ2/Q3 current by 0.65 ± 0.12-fold, which was significantly less than the control (1.68 ± 0.06, *, p < 0.05).