Fig. 5: Osmotic stress promotes PROFILIN mediated actin-reorganization, maintaining cell and tissue integrity upon osmotic stress.

a Actin localization in the Arabidopsis primary root elongation zone of wild-type roots on control conditions (CTL), transferred to 50 mM NaCl, 140 mM NaCl or 300 mM mannitol for 16 h (left to right). Images from treatments correspond to cells actively elongating at the time of transfer. Actin filaments were visualized with the actin binding domain of the FIMBRIN protein tagged with green fluorescent protein (ABD2:GFP). Scale bar = 50 µm. b Quantification of skewness and angle of actin filaments relative to the cell radial axis. Each point represents a cell that was actively elongating when roots were transferred. n > 25 cells per condition, taken from three independent experiments. Center lines show medians and box limits indicate the 25th and 75th percentiles. Whiskers represent minima and maxima, (*) P values are from a one-way ANOVA test between control condition and treatments, p < 0.05. See Source Data. c Actin localization in the Arabidopsis root elongation zone of prf5-1 roots on control conditions (CTL), transferred to 50 mM NaCl, 140 mM NaCl, or 300 mM mannitol for 16 h (left to right). Images from treatments correspond to cells actively elongating at the time of transfer. Actin filaments are visualized with ABD2:GFP. Due to silencing of the reporter and a loss of cell viability, some cells within the field of view are not marked by GFP expression. Scale bar = 50 µm. See Source Data. d Quantification of skewness and angle of actin filaments relative to the cell radial axis. Each point represents a cell that was actively elongating when roots were transferred. n = 25 cells per condition, taken from three independent experiments. Center lines show medians and box limits indicate the 25th and 75th percentiles. Whiskers represent minima and maxima, (*) P values are from a one-way ANOVA test between control condition and treatments, p < 0.05. See Source Data. e 16 colors LUT confocal images of primary root elongation zone expressing GCamp6, and prf5-1, GCamp6 in control conditions (0) or upon transfer to 50 mM NaCl for ½ h and 1 h. Images represent z-projection in each timepoint. Scale bar = 50 µm. f Hourly average number of [Ca²⁺] spikes in wild-type (GCamp6) cells and prf5-1 with introgressed GCamp6 (prf5-1;GCamp6) treated 50 mM NaCl. n = 6 roots, taken from three independent experiments. Center lines show medians and box limits indicate the 25th and 75th percentiles. Whiskers represent minima and maxima, * indicates significance, one-way ANOVA test between genotypes 1 h after transfer to 50 mM NaCl, p = 0.0007179. See Source Data. g Confocal images of 5-day old primary root expressing membrane marker LTI-YFP in wild-type plants and prf5-1 mutants. 250 and 500 mM Mannitol represent the plasmolysis treatments performed for 60 min. Control roots were incubated in liquid MS media lacking Mannitol. Scale bar = 50 µm. h Quantification of protoplast shrinkage in wild-type (Col-0) and prf5-1 mutants. Percentage of shrinkage was calculated based on the protoplast area (LTI-YFP signal) and the cell wall area (bright field channel) (see Material and Methods). n = 37 cells, taken from three independent experiments. Center lines show medians and box limits indicate the 25th and 75th percentiles. Whiskers represent minima and maxima, * indicates significance, one-way ANOVA test between different genotypes or treatments within the same plasmolysis regime, p < 0.001. See Source Data. i Cartoon illustrating actin filaments remodeling upon osmotic stress to promote tissue integrity.