Figure 6.

Effects of the photosynthesis inhibitor DCMU on the light responses of guard cells in whole leaves. Mature leaves harvested from dark-adapted plants were pretreated with 10 µm DCMU dissolved in 0.1% (v/v) DMSO (+DCMU) or 0.1% (v/v) DMSO alone (–DCMU) for 30 min in the dark before light illumination. A, Immunohistochemical detection of the phosphorylation of PM H⁺-ATPase in guard cells using anti-pThr antiserum. Pretreated leaves were illuminated with red light (600 µmol m^–2 s^–1) for 30 min (R30) followed by blue light (5 µmol m^–2 s^–1) superimposed on red light for 2.5 min (R + B2.5) or kept in the dark (D30). Typical fluorescence and bright-field images (top) and quantification of the fluorescence intensities (bottom) are shown. Arrowheads indicate guard cells. Data represent means of relative values from five independent measurements with sd. Daggers indicate that the mean is statistically significantly higher than D30 of –DCMU set to 1. N.S., Not significant (one-tailed one-sample Student’s t test: ^†, P < 0.05; ^††, P < 0.001; and N.S., P > 0.58). Asterisks indicate that the mean of R + B2.5 is statistically significantly higher than that of R30 within each treatment and that the mean of R + B2.5 of +DCMU is statistically significantly lower than that of –DCMU (one-tailed Student’s t test: *, P < 0.05; **, P < 0.01; and ***, P < 0.005). Bars = 50 µm. B, Measurement of light-induced stomatal opening under DCMU treatment. Pretreated leaves were illuminated with red light (R60), both red and blue light (RB60), or kept in the dark (D60) for 60 min. Light intensities were the same as in A. Data represent means of representative values from four independent measurements with sd. Different letters indicate statistically significant differences among means (Tukey’s test: P < 0.05).