Extended Data Fig. 9 |. BSL Requires BIN2 and YDA to Regulate Stomatal Development in Arabidopsis.

a, Overexpression phenotypes of the TMM promoter driven BSL1 (green) in 5-day adaxial cotyledon epidermis. Right, qPCR data show relative expression levels of BSL1 in two independent overexpression lines used in this study. Total RNAs were extracted from 3-day-old seedlings. Gene expression levels were normalized by ACTIN2 and relative expression levels of BSL1 were compared with the values in the wild type. Data are presented as mean ± SD. n = three independent experiments. Primers used were listed in Supplementary Table 2. b, Stomatal phenotypes in 5-day adaxial cotyledon epidermis overexpressing phosphatase-dead BSL1^D584N (green) in wild-type (left) and overexpressing wild-type BSL1 (red, right) in the basl-2 mutant (right), both driven by the TMM promoter. Note the stomatal phenotype generated by BSL1^D584N resembled that observed for bsl-quad mutants, suggesting a dominant negative effect of BSL1^D584N. c, Overexpression of BSL1 (green) driven by the TMM promoter in the loss of function yda-3 mutant did not alter the stomatal phenotypes in yda-3. d, Overexpression of constitutively active YDA (YDA^CA, green) driven by the SPCH promoter in the wild-type (left) and in bsl-quad mutants (right). The activities of YDA^CA suppressed the loss-of-function phenotypes in bsl-quad. e, Overexpression of BSL1 (green) driven by the TMM promoter (BSL1 ++) in the loss-of-function mutants bin2–3;bil1;bil2 (bin). bin mutations appeared to be epistatic. Cell outlines were stained with Propidium Iodide (magenta). f, Stomatal phenotype in 5-dpg adaxial cotyledon epidermis of Ws-2 (background of bin2). Right, quantification of Stomatal Index (# stomata relative to # total cells) of the designated genotypes. Box plot shows first and third quartiles, median (line) and mean (cross). n, number of cotyledons used for quantification. One-way ANOVA with Tukey’s post hoc test were used to compare with the wild-type Col-0. n.s., not significant. *** P < 0.0001. Scale bars in (a-e), 50 μm.