Fig. 2: AtEB1 proteins function in the redistribution of KAT1-mNeonGreen to the PM and affect intracellular K⁺ content.

a Distribution of mNeonGreen-tagged KAT1 (KAT1-mNeonGreen) in guard cells from wild-type (Col-0) and eb1a 1b 1c mutant plants before and after light treatment. Light stimulus increased the redistribution of KAT1-mNeonGreen to the PM in wild-type guard cells, whereas this process was dramatically delayed in eb1a 1b 1c cells. Z-stack images (0.5-μm step size) were acquired, and the max-intensity projection is shown. Total four independent experiments were performed and obtained similar results. Bars = 5 µm. b The ratio of mean KAT1-mNeonGreen fluorescence intensity between the PM and cytosol (excluding chloroplast auto-fluorescence) was measured in guard cells from wild-type and eb1a 1b 1c mutant plants as shown in a. c Representative pseudo-coloured images of intracellular K⁺ levels in guard cells expressing lc-LysM GEPII1.0 during a light-induced stomatal opening process. Total of five independent experiments were performed and obtained similar results. Bars = 5 µm. d Quantitative analysis of light-induced intracellular K⁺ dynamics from wild-type and eb1a 1b 1c mutant guard cells based on FRET ratios (FRET/CFP). Data in b and d are presented as boxplots. The whiskers indicate the maximum/minimum values and the centre line within the box reflects the median value and the boundary lines of box indicate the 1^st quartile (upper) and the 3^rd quartile (lower) value of the dataset. The data were analysed using RStudio software with the agricolae package. Different letters indicate significant differences analysed by one-way ANOVA with Duncan’s new multiple range test, P < 0.05. The “n” below each box indicates the number of analysed guard cells.