Fig. 6: TOT3 impinges brassinosteroid signalling pathway.

a Representative western blot (of 3 biological replicates) of BES1 phosphorylation status in 4-day-old Col-0 and tot3-2 seedlings grown on mock (DMSO) or 100 nM brassinolide (BL) at 28 °C under short-day conditions. BES1 was detected with anti-BES1 antibody. BES1-P, phosphorylated BES1; BES1, dephosphorylated BES1. Detection of TUBULIN (TUB) was used as a loading control. b Hypocotyl length of 5-day old tot3-2 and Col-0 wild-type plants grown at 21 and 28 °C in short-day conditions on medium containing DMSO or 50 nM brassinolide. c, d Hypocotyl length of 4-day old tot3-2, bzr1-1D, tot3-2 bzr1-1D and Col-0 wild-type seedlings grown at 21 and 28 °C in darkness on MS/2 medium containing DMSO (c) or 1 µM brassinazole (d). e Relative expression of BZR1 target genes PRE6, PAR1 and PAR2 in Col-0 and tot3-2 plants grown at control (21 °C) and warm temperature (28 °C) under short-day conditions. f Relative expression of BZR1 target genes PRE6, PAR1 and PAR2 in Col-0 wild-type and tot3-2 plants grown at control (21 °C) and warm temperature (28 °C) under dark conditions. Box plots show median with Tukey-based whiskers and outliers (b, c, d). The number of individually measured seedlings (n) is indicated above the X-axis (b, c, d). Bar diagram shows mean of 3 biological replicates (individual dots) with standard error of the mean (e, f). Letters indicate significant differences based on two-way ANOVA and Tukey’s test (p < 0.01); n.s: not significant. The p-value for the interaction (genotype × temperature or genotype × treatment) is shown at the top.