Fig 1.
Primary root length, number of lateral roots, lateral roots length, root hair length, and root hair density.
A) Dose-response curves of PRL (primary root length), B) NLR (number of lateral roots), C) LRL (lateral roots length), D) RHL (root hair length), and E) RHD (root hair density) of 250 μM farnesene-treated Arabidopsis seedlings for 14 days. Data are expressed as percentage of the control. Different letters indicate significant differences among treatments at P ≤ 0.05. N = 5.
Fig 2.
Morphology of Arabidopsis root tips.
Arabidopsis root tips grown on agar medium (A) Control root tip with root hairs (B) Root tips treated with 250 μM farnesene. Note the absence of root hairs and the anticlockwise torsion of the cell lines (left-handed). On the bottom is reported a panoramic of young Arabidopsis seedlings grown on agar medium untreated (C) and treated with 250 μM farnesene (D). Note the anisotropic growth of the root and the apparent loss of gravitropic response.
Fig 3.
Bioassay on root gravitropism.
Effect of farnesene on the time course gravitropic curvature of Arabidopsis thaliana primary roots. After 90° rotation, images were taken after 6, 12, 24, 48 and 96 hours. Data were analyzed through t-test (P ≤ 0.05). Circular graphs indicate the distribution of the angle of root curvature. * = (P ≤ 0.05), ** = (P ≤ 0.01), *** = (P ≤ 0.001). N = 10.
Fig 4.
Arabidopsis seedlings grown in taxol and/or farnesene for 14 days.
A) Seedlings at time zero, when treatment started; B) Untreated (control); C) 250 μM farnesene; D) 250 μM farnesene + 0.5 μM taxol. Circular graphs indicate the angle of root curvature. * = (P ≤ 0.05), ** = (P ≤ 0.01), *** = (P ≤ 0.001). Images magnification 20X. N = 5.
Table 1.
Effects of 250 μM farnesene on the ultra-structure of 7 and 14 days treated roots.
Cell ultra-structure and changes in the organelles were identified analyzing transmission electron microscopic images of farnesene-treated and untreated roots. A number of 200 images were analyzed for this table.
Table 2.
Quantification of changes detected in the cell structure and the organelles of 250 μM farnesene-treated and untreated (0 μM) Arabidopsis roots.
A number of 100 TEM images was used for the quantification of each parameter.
Fig 5.
TEM images of farnesene-treated and untreated Arabidopsis meristems.
TEM images of the apical meristem of untreated (A, D, G, J, M, P, S), and 7 days (B, E, H, K, N, Q, T) and 14 days (C, F, I, L, O, R, U) farnesene-treated Arabidopsis roots: A) Nucleus of a stele cell; B) Binucleated cell of a stele cell; C) High presence of polynucleated cells; D) Epidermal cells with normal cell division and phragmoplast formation; E) Epidermal cells with high presence of cell wall deposits on cell corners; F) Protodermal cells with abnormal shape and swollen cell walls; G) Normal cell corner without deposits; H and I) High presence of cell wall deposits; J) Plasmodesmata; K and L) High presence of incomplete plasmodesmata; M) Regular Golgi apparatus; N) Golgi apparatus with massive production of vesicles; O) High presence of Golgi apparatus and vesicles; P) Normally vacuolated cortex and endodermic cells; Q and R) Endodermal cell: increase in size and number of the vacuoles; S) Mitochondria with regular morphology and a good number of cristae; T) Broken mitochondria, High number of Golgi vesicles; U) Increased number of mitochondria characterized by irregular shape and swollen/translucent stroma. Nucleus (N), vacuole (V), cell wall (CW), mitochondria (M), binucleated cell (BC), phragmoplast (F), cell wall deposit (CWD), cell corner (CC), plasmodesmata (P), endoplasmic reticulum (ER), incomplete plasmodesmata (IP), Golgi apparatus (G), Golgi vesicles (GV), uncompleted phragmoplast (UF).
Fig 6.
TEM images of cortical microtubules.
TEM images of cortical microtubules (transversal section) in cells of Arabidopsis roots grown in 250 μM farnesene for 7 and 14 days. A) Cell of 7 days control roots; B) Cell of 14 days control roots; C) Cell of 7 days farnesene-treated roots; D) Cell of 14 days farnesene-treated roots. Black arrows indicate transversally cut microtubules.
Fig 7.
Microtubule immunostaining of farnesene and taxol treated and untreated Arabidopsis roots.
Figure shows (A) control cells after 7; and (B)14 days; (C) cells treated with 250 μM farnesene for 7 and (D)14 days; (E) cells treated with 250 μM farnesene + 0.5 μM taxol for 7 and (F)14 days. Images were acquired by confocal microscopy (63X immersion objective). (G) Fluorescence quantification of immunolabeled microtubules of roots treated for 7 and 14 days: control (C); farnesene 250 μM (F); taxol 0.5 μM (T). Data are expressed as arbitrary units (AU). Different letters between the bars indicate significant differences at P ≤ 0.05 (LSD test). N = 5. Fluorescence was quantified using the open source software ImageJ.
Fig 8.
Auxin and ethylene contents of farnesene-treated and untreated seedlings.
A) Auxin concentration; B) Ethylene emission after incubation in distilled water (real level); C) Ethylene emission after incubation in ACC (ACC addition leads to obtain maximal ethylene emission) in A. thaliana roots after 7 and 14 days treated with 250 μM farnesene. Data are expressed as percentage of the control. * = (P ≤ 0.05), ** = (P ≤ 0.01), *** = (P ≤ 0.001). N = 6.
Fig 9.
In situ hydrogen peroxide and superoxide localization in farnesene-treated and untreated roots.
In situ hydrogen peroxide (A-C) and superoxide (D-F) localization in roots of Arabidopsis treated with 250 μM farnesene for 7 and 14 days. Control (a and d), 7 days (b and e), 14 days (c and f). (G) NO emission (expressed as arbitrary units) from Arabidopsis roots treated with 250 μM farnesene for 7 and 14 days. * = (P ≤ 0.05), ** = (P ≤ 0.01), *** = (P ≤ 0.001). Image magnification 12X. N = 3.