Cortical microtubules and fusicoccin response in clustered stomatal guard cells induced by sucrose solution immersion

<p>We previously found that sucrose solution immersion treatment permitted ectopic guard cell differentiation, resulting in clustered stomatal guard cells. Using this system, we examined the effects of sucrose solution-induced stomatal clustering on guard cell cortical microtubules and the stomatal response to fusicoccin. Confocal observation revealed that the radial orientation of cortical microtubules was largely maintained in clustered guard cells. Outward movement of cortical microtubule plus-ends was also kept in the clustered guard cells. Fusicoccin treatment induced stomatal opening in both spaced and clustered stomata, although sucrose solution-treated guard cells had lower stomatal apertures. These results suggested that immersion treatment with sucrose solution perturbed the one-cell spacing of stomata but not the cortical microtubule organization required to open stomatal pores.</p

Breaking of Plant Stomatal One-Cell-Spacing Rule by Sugar Solution Immersion

<div><p>The spatial distribution of plant stomata is a model system to study epidermal cell pattern formation. Molecular genetic approaches have identified several key genes required for stomatal distribution patterning, but environmental conditions that perturb the stomatal spacing distribution have not yet been identified. We found that immersing hydroponic cultures in 1–5% sucrose solution induced abnormally clustered stomata in the cotyledons of Arabidopsis seedlings. Clustered stomata were also induced by treatment with glucose or fructose solution but not by mannitol solution, suggesting that osmotic stress was not a cause of the disturbed stomatal patterns. Stomatal lineage cell-specific enhancer trap lines revealed that the sugar solution treatment led to ectopic expression of stomatal lineage cell-specific genes in non-stomatal lineage cells. Aniline blue staining also showed that there was reduced deposition of callose, a plant cell wall component, in new cell walls during formation of stomatal precursor cells (meristemoids). These results suggested that the immersion treatment with sugar solution permitted ectopic guard cell differentiation through dysfunction of the cell wall dividing stomatal- and non-stomatal lineage cells. Our simple induction system for clustered stomata provides a suitable tool for further studies to investigate the one-cell-spacing rule during plant stomatal development.</p></div

Radiocesium Distribution in Bamboo Shoots after the Fukushima Nuclear Accident

<div><p>The distribution of radiocesium was examined in bamboo shoots, <i>Phyllostachys pubescens</i>, collected from 10 sites located some 41 to 1140 km from the Fukushima Daiichi nuclear power plant, Japan, in the Spring of 2012, 1 year after the Fukushima nuclear accident. Maximum activity concentrations for radiocesium ¹³⁴Cs and ¹³⁷Cs in the edible bamboo shoot parts, 41 km away from the Fukushima Daiichi plant, were in excess of 15.3 and 21.8 kBq/kg (dry weight basis; 1.34 and 1.92 kBq/kg, fresh weight), respectively. In the radiocesium-contaminated samples, the radiocesium activities were higher in the inner tip parts, including the upper edible parts and the apical culm sheath, than in the hardened culm sheath and underground basal parts. The radiocesium/potassium ratios also tended to be higher in the inner tip parts. The radiocesium activities increased with bamboo shoot length in another bamboo species, <i>Phyllostachys bambusoides</i>, suggesting that radiocesium accumulated in the inner tip parts during growth of the shoots.</p></div

Aniline blue staining of cotyledon epidermal cells.

<p>Four day-old cotyledons immersed in sugar-free (A) or 3% sucrose (B) solutions were stained with 0.02% aniline blue for 1 week. Representative images from 24 (sugar-free) and 38 (3% sucrose) independent seedlings were shown. Note that aniline blue fluorescence was clearly detected in new cell walls forming in meristemoids immersed in sucrose-free solutions but not in 3% sucrose solutions. Scale bars  = 10 μm.</p

List of sampling bamboo shoots and their Cs-134 and -137 concentrations in the upper edible part.

<p>*USS: under surface soil.</p

Percentage of stomata in each cluster size class.

<p>Abaxial cotyledons from 12- to 15 day-old seedlings grown in sugar-free, 1, 3 or 5% sucrose (A), and 3% glucose, fructose or mannitol (B) solutions were subjected to quantitative analysis. Data are mean values of 20–68 independent observations. Significance with sugar-free conditions was determined using Mann–Whitney's U-test. <i>p</i>-value *<0.0001. Total number of stomata counted: <i>n</i> = 281–1843.</p

Radiocesium contamination in bamboo shoots of <i>Phyllostachys pubescens</i> with an outcrop length of 3.5 m.

<p>(a) The separated tall bamboo shoot collected in Kashiwa in Chiba Prefecture (195 km from Fukushima Daiichi). The stripped bamboo shoots were cut at 30 cm intervals from the tip, separating the upper edible part, and the middle parts 1, 2 and 3. Scale bar indicates 30 cm. (b, c). The radioactive concentrations of radiocesium, ¹³⁴Cs (b) and ¹³⁷Cs (c), in each part of the bamboo shoots shown in (a). Error bars show one standard deviation.</p

Relationship between outcrop lengths and the radiocesium concentrations in bamboo shoots of <i>Phyllostachys bambusoides</i> Sieb. Et Zucc.

<p>(a) The collected <i>Phyllostachys bambusoides</i> bamboo shoots. Scale bar indicates 10 cm. (b) The scatter plot of the radioactive concentrations of radiocesium in tip parts and the outcrop length. (c, d) The radioactive concentrations of radiocesium, ¹³⁴Cs (c) and ¹³⁷Cs (d), in each part of the bamboo shoots shown in (a). Error bars show measurement deviation.</p

Sampling and fractionation of bamboo shoots.

<p>(a) Map of Japan showing the location of the twelve sampling sites and the Fukushima Daiichi nuclear power plant. (b) Schematic workflow of the bamboo shoot fractionation. The basal part was cut off (black down-arrow), and about 10–15 culm sheaths were stripped (black right-arrow). The culm sheaths were cut into the upper and lower parts. The stripped bamboo shoot was separated into the apical culm sheath and the upper and lower edible parts (white arrows).</p

Comparison of radiocesium and potassium in each part of the bamboo shoots of <i>Phyllostachys pubescens</i>.

<p>(a, b) Scatter plots of the radioactive concentrations of radiocesium, ¹³⁴Cs (a) and ¹³⁷Cs (b), and the potassium concentrations. UEP, LEP, BP, ACS, UCS, LCS indicate the upper edible part, lower edible part, basal part, apical culm sheath, upper culm sheath and lower culm sheath, respectively. (c, d) The mean ratio of the radioactive concentrations of radiocesium, ¹³⁴Cs (c) and ¹³⁷Cs (d), to the potassium concentrations. Error bars show the standard deviation for three separate bamboo shoots in Tsukubamirai as shown in (a, b). Significance was determined using Welch's two-sample test. <i>p</i>-value *<0.01, **<0.003, ***<0.001.</p