Figure 2.

Simultaneous cytosolic and ER Ca²⁺ analyses in root-tip cells treated with the P-Type IIA inhibitor CPA. A, Green: ER-GCaMP6-210 fluorescence, magenta: R-GECO1 fluorescence, and overlay. Scale bar 50 µm. B, Examples of false-color images illustrate R-GECO1 (magenta) and ER-GCaMP6-210 (green) of the selected cell (dashed rectangle in A) in root tips of seedlings expressing simultaneously the two Ca²⁺ sensors at steady-state and during the Ca² treatment with 25 µM CPA for 3 min. C, R-GECO1 and ER-GCaMP6-210 normalized fluorescence changes of the selected cell over the time acquired under continuous perfusion and treated with 25 µM CPA for 3 min, as indicated by the black box on the x-axis. C′, R-GECO1 and ER-GCaMP6-210 normalized fluorescence of the selected cell over the time acquired under continuous perfusion and treated with DMSO (the CPA solvent) as a control for 3 min, as indicated by the black box on the x-axis. D, same as panel (C) but x-axis, y-axis scales, and ranges adjusted. The double arrow in (D) indicates the delay time quantified in (E). E, Mean delay of the fluorescence increase of the R-GECO1 following the ER-GCaMP6-210 fluorescence decrease. F, The maximal peak of R-GECO1 and the minimal level of ER-GCaMP6-210 fluorescence signals for the selected cells after CPA administration. n = 5. Error bars = sd.