Fig. 4

PAMP significantly altered cytoskeletal architecture and dynamics as revealed by dual-color VAEM. To investigate whether the two cytoskeletal structures interact coordinately, seedlings expressing green fluorescent protein (GFP)-F-actin binding domain of fimbrin2 (fABD2) and mCherry-α-tubulin 5 isoform (TUA5) were generated by crossing. LSCM and VAEM were carried out on the seedlings respectively. a AFs and MTs in mock-treated epidermal cells from the wild-type cells, arrows indicate actin fragments reside in transient coincidence with MTs; b Selected frames from a time series of a GFP-fABD2 and mCherry-TUA5 dual-labeled line; c AFs and MTs in the wild-type epidermal cells. d to e AFs appeared to be more abundant after flg22 and chitin treatment, while the transverse MT organization gradually changed to longitudinal fragments. Epidermal cells treated with 1 μM flg22 or 10 μM chitin had a significant increase in AF abundance compared with mock-treated seedlings. f To address the specificity for the elicitor-triggered reorganization of cytoskeletal components as above, 50 μM isoxaben, a cellulose synthesis inhibitor, was applied to treat the seedlings. The MTs were dramatically disordered following incubation while AFs did not show significant changes in density and intensity. g Average filament density (percentage of occupancy) analysis was performed on images collected from epidermal cells in the cotyledons. When compared with mock-treated cells, the AF density was significantly increased after 1 μM flg22 and 10 μM chitin treatments; in contrast, MT density decreased significantly after chitin and flg22 treatment, while isoxaben treatment significantly decrease MT density without affecting AF density; The extent of filament bundling, or skewness, was measured. No significant differences were observed among treatments. Values given were mean ± SE. (n = 100 cells from 10 cotyledons per treatment; ** P < 0.001; ND, no significant difference; Student’s t-test). Bars = 10 μm. Interval between frames = 200 ms, “0 s” referred to the time point at 5 min after corresponding treatment