Extended Data Fig. 10: Basal [Ca²⁺]_cyt level and stimulus-induced [Ca²⁺]_cyt elevations are largely comparable between WT and ica1/2/3/4 mutants.

a-d, [Ca²⁺]_cyt responses to NaCl and mannitol in root epidermal and cortical cells of plants expressing the FRET-based Ca²⁺ sensor YC3.6. Measurements were performed in root epidermal and cortical cells of WT^YC3.6 and ica1/2/3/4^YC3.6 seedlings. Roots were imaged every 2 s. Time course of YC3.6 emission ratio changes in response to local application of 200 mM NaCl (a). Basal (pre-stimulus at 12 s) and peak YC3.6 emission ratios derived from experiments as in (a) were shown in (b). Time course of YC3.6 emission ratio changes in response to local application of 600 mM mannitol (c). Basal (pre-stimulus) and peak YC3.6 emission ratios derived from experiments as in (c) were shown in (d). e-g, High salt-induced local Ca²⁺ signals were similar in WT^GCaMP6s and ia1/2/3/4^GCaMP6s roots. Measurements were performed in WT^GCaMP6s and ica1/2/3/4^GCaMP6s seedlings. Roots were imaged every 1 s following local application of 200 mM NaCl. Representative GCaMP6s fluorescence images of roots from WT^GCaMP6s and ica1/2/3/4^GCaMP6s before treatment (0 s), and at 40 s and 180 s after application of 200 mM NaCl (e). Red circles indicate the local region of interest (ROI) analyzed in (f). Scale bars, 100 µm. Time course of GCaMP6s fluorescence changes at the local ROI (f). Peak GCaMP6s fluorescence changes from (f) were shown in (g). h-j, High salt-induced propagation of Ca²⁺ waves were similar in WT^GCaMP6s and ia1/2/3/4^GCaMP6s roots. Representative GCaMP6s fluorescence images (as in e) with the red circle marking the distal ROI for wave analysis (h). Time course of GCaMP6s fluorescence changes in the distal ROI (i). Peak GCaMP6s fluorescence changes from (i) were shown in (j). Data are mean ± s.e.m. (a,c,f,i), mean ±s.d. (b,d,g,j). P values were determined using two-tailed Student’s t-tests. Experiments were repeated three times with similar results.

Source data