Aluminum Induces Distinct Changes in the Metabolism of Reactive Oxygen and Nitrogen Species in the Roots of Two Wheat Genotypes with Different Aluminum Resistance

Aluminum (Al) toxicity in acid soils is a primary factor limiting plant growth and crop yield worldwide. Considerable genotypic variation in resistance to Al toxicity has been observed in many crop species. In wheat (<i>Triticum aestivum</i> L.), Al phytotoxicity is a complex phenomenon involving multiple physiological mechanisms which are yet to be fully characterized. To elucidate the physiological and molecular basis of Al toxicity in wheat, we performed a detailed analysis of reactive oxygen species (ROS) and reactive nitrogen species (RNS) under Al stress in one Al-tolerant (Jian-864) and one Al-sensitive (Yang-5) genotype. We found Al induced a significant reduction in root growth with the magnitude of reduction always being greater in Yang-5 than in Jian-864. These reductions were accompanied by significant differences in changes in antioxidant enzymes and the nitric oxide (NO) metabolism in these two genotypes. In the Al-sensitive genotype Yang-5, Al induced a significant increase in ROS, NO, peroxynitrite (ONOO^–) and activities of NADPH oxidase, peroxidase, and S-nitrosoglutathione reductase (GSNOR). A concomitant reduction in glutathione and increase in S-nitrosoglutathione contents was also observed in Yang-5. In contrast, the Al-tolerant genotype Jian-864 showed lower levels of lipid peroxidation, ROS and RNS accumulation, which was likely achieved through the adjustment of its antioxidant defense system to maintain redox state of the cell. These results indicate that Al stress affected redox state and NO metabolism and caused nitro-oxidative stress in wheat. Our findings suggest that these molecules could be useful parameters for evaluating physiological conditions in wheat and other crop species under adverse conditions

Cd and Zn tolerance of yeast cells expressing <i>SaMT2</i>.

<p>The <i>Saccharomyces cerevisiae</i> BY4741, <i>Δycf1</i> and <i>Δzrc1</i> yeast cells harboring pDR195 (vector control) or pDR195-<i>SaMT2</i> were grown in liquid SD selective medium. Cultures were adjusted to OD_600nm of 0.1 and serially 10-fold diluted in water. 10 µl aliquots of each dilution were spotted either on SD selective plates or on plates with 30 µM CdCl₂ or 5 mM ZnSO₄. After 3 days of incubation at 30°C, plates were photographed. CK represents the control group.</p

Cd and Zn concentration in <i>Δycf1</i> and <i>Δzrc1</i> yeast cells expressing <i>SaMT2</i>.

<p>The yeast transformants containing pDR195 or pDR195-<i>SaMT2</i> were grown in liquid SD selective medium with 30 µM CdCl₂ and 100 µM ZnSO₄ for <i>Δycf1 and </i><i>Δzrc1</i>, respectively. Cells were incubated at 30°C for 48 h and metal contents were measured by ICP-MS. Results are averages (±S.E.) from three independent experiments done with four different colonies. The ‘*’ symbol indicates the mean values were significantly different at p<0.05 (Tukey’s test).</p

Sequence alignment and phylogenic analysis of <i>SaMT2</i> with other MTs.

<p>(A) The deduced amino acid sequences encoded by <i>SaMT2</i> were aligned with MTs from <i>Arabidopsis thaliana</i>, <i>Noccaea caerulescens</i> and <i>Solanum nigrum</i>. The cysteine-rich domains are boxed. (B) The phylogenic tree of <i>SaMT2</i> and MTs from Arabidopsis and rice.</p

Metal tolerance analysis of transgenic tobacco plants over-expressing <i>SaMT2</i>.

<p>The figure shows the effect of 200 µM ZnSO₄ or 100 µM CdCl₂ on the growth of WT and transgenic plants on B5 medium. CK represents the control group.</p

Cd and Zn concentrations in wild type amd transgenic tobacco lines overexpressing <i>SaMT2</i>.

<p>Three independent <i>SaMT2</i> over-expressing lines and wild-type tobacco were grown in nutrient solution containing 50 µM CdCl₂, 100 µM ZnSO₄ for 1 week. A: Cd concentration in roots, B: Cd concentrantion in shoots, C: Zn concentration in roots, D: Zn concentration in shoots. Results are means ± S.E. (n = 3). Different letter indicate the mean values were significantly different from WT tobacco determined by Tukey’s test (p<0.05).</p

Relative root growth of transgenic tobacco plants.

<p>The relative root growth of WT and transgenic tobacco plants under Cd (A) and Zn (B) treatments. Different letters above the columns indicate a significant difference among different plant lines (p<0.05, Tukey’s test).</p

The expression level of <i>SaMT2</i> in <i>Sedum alfredii</i>.

<p>The transcript level of <i>SaMT2</i> induced by Cd and Zn treatments. The different letters above the columns indicate the significant difference between the treatments (p<0.05, Tukey’s test). CK represents the control group.</p

µ-XRF elemental maps of early stage rice seedlings after seed germination for 48

<p> <b>h.</b> The orientation of the individual µ-XRF elemental maps and the color-merged image (upper right) is the same as that of the green rectangle around a portion of the labeled image of the germinating seed. Refer to the legend for <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0057360#pone-0057360-g004" target="_blank">Figure 4</a> for additional details.</p

Elemental concentrations of rice grains, hull, brown rice, bran, and polished rice.

<p>All concentrations are expressed as mg kg⁻¹ DW. Data points and error bars represent means and SEs of four replicates.</p