Fig 7. Role of QKY in isoxaben-induced CBI responses.

(A) Quantification of H₂CDFDA signal indicating ROS accumulation in root tips of six-day-old Col-0 and qky-11 seedlings exposed to mock or 600 nM isoxaben for the indicated time. Note the high signal in qky-11 after a 30- or 60-minutes exposure to isoxaben. Box and whisker plots are shown. Whisker ends mark the minimum and maximum of all the data. 10 ≤ n ≤ 14. Asterisks represent adjusted P values (_* P < 0.04, _**** P < 0.0001; ns: not significant; one-way ANOVA followed by Tukey’s multiple comparison tests). No statistically significant differences were measured within or between genotypes at the zero minute time point of mock or isoxaben treatment or between wild-type and qky-11 mock treatment at the 30 and 60 minutes time points (P > 0.8557; one-way ANOVA followed by Tukey’s multiple comparison tests). Experiments were performed three times with similar results. (B) Gene expression levels of several CBI marker genes by qPCR upon exposure of seven-day-old seedlings to 600 nM isoxaben for eight hours. The results from three biological replicates are shown. Marker genes and genotypes are indicated. Mean ± SD is presented. The experiment was repeated three times with similar results. (C) Phloroglucinol signal strength indicating lignin accumulation in roots of six-day-old seedlings exposed to 600 nM isoxaben for 12 hours. Genotypes: Col, qky-11 (Col), Ler, and sub-1 (Ler). (D) Quantification of the results depicted in (C). Genotypes are indicated. Box and whisker plots are shown. Whisker ends mark the minimum and maximum of all the data. 15 ≤ n ≤ 19. Asterisks represent statistical significance (_* P < 0.04, _** P < 0.01; unpaired t test, two-tailed P values). The experiment was performed three times with similar results. (E) Confocal micrographs show cotyledons of seven-day-old Ler, qky-8 (Ler), Col, and qky-11 (Col) seedlings treated with mock or 600 nM isoxaben for 24 hours. Aniline blue fluorescence signal strength indicates callose accumulation. (F) Quantification of the results depicted in (E). Left panel shows results obtained from qky-8 mutants in Ler background. Right panel indicates results obtained from qky-11 mutants in Col background. Box and whisker plots are shown. Whisker ends mark the minimum and maximum of all the data. 5 ≤ n ≤ 11. Asterisks represent adjusted P values (_* P < 0.04, _*** P < 0.0005, _**** P < 0.0001; one-way ANOVA followed by Tukey’s multiple comparison tests). The experiment was performed three times with similar results. Abbreviation: ISX, isoxaben. Scale bars: (C) 0.1 mm; (E) 0.2 mm.