Fig. 2: PRAF proteins are required for stomatal ACD and BASL polarization. | Nature Communications

Fig. 2: PRAF proteins are required for stomatal ACD and BASL polarization.

From: Connected function of PRAF/RLD and GNOM in membrane trafficking controls intrinsic cell polarity in plants

Fig. 2

a Confocal images show abnormal stomatal division and differentiation in praf mutants. Five-day-old adaxial side cotyledon epidermis of the designated genotypes was examined. Cell outlines were visualized with Propidium Iodide (PI) staining and images were converted to black/white. Stomatal lineage cells were manually traced and highlighted by blue and abnormal guard cells were highlighted in purple. Data represent results of three independent experiments. Scale bars, 25 μm. b–c Quantification of stomata lineage index (b, ratio of # stomatal lineage cells/# total epidermal cells) and divisional asymmetry (c, ratio of cell sizes) for the genotypes shown in (a). Box plots show first and third quartile (box), median (line) and mean (cross). The parameters are applicable to all quantification data presented as box plots in this study. n, # cotyledons counted. Student’s unpaired t tests were used for comparison. Two-sided P values are <0.0001 (for wild type vs. praf4c;5c;8c;9c and wild type vs. praf4t;5c;8c;9c), 0.0003 (for basl-2 vs. praf4t;5c;8c;9c), 0.0027 (for basl-2 vs. basl-2; praf4c;5c;8c;9c), and 0.0489 (for praf4c;5c;8c;9c vs. basl-2; praf4c;5c;8c;9c) in (b); and <0.0001 (for wild type vs. praf4c;5c;8c;9c and wild type vs. praf4t;5c;8c;9c), 0.7966 (for basl-2 vs. praf4t;5c;8c;9c), 0.1664 (for basl-2 vs. basl-2;praf4c;5c;8c;9c), and 0.3407 (for praf4c;5c;8c;9c vs. basl-2; praf4c;5c;8c;9c) in (c). n.s. not significant; *P < 0.05; **P < 0.005; ***P < 0.001; ****P < 0.0001. d–f Confocal images show disturbed GFP-BASL (green) polarization in praf4t;5c;8c;9c mutants (e) or upon the PRAF-interacting FxxFxF motif mutated (BASL_3F > 3K) (f). Data represent results of three independent experiments. Arrows in (d) indicate typical BASL polarization. White brackets in (f) show stomatal defects (clusters) typical for a basl-2 mutant. PI staining was used to highlight cell outlines (magenta). Scale bars, 10 μm. (z), z-stacked confocal image. Quantification of BASL polarization in (d–f) is shown in Supplementary Fig. 3d.

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