Nitric Oxide Enhancing Resistance to PEG-Induced Water Deficiency is Associated with the Primary Photosynthesis Reaction in Triticum aestivum L

Abstract

Photosynthesis is affected by water-deficiency (WD) stress, and nitric oxide (NO) is a free radical that participates in the photosynthesis process. Previous studies have suggested that NO regulates excitation-energy distribution of photosynthesis under WD stress. Here, quantitative phosphoproteomic profiling was conducted using iTRAQ. Differentially phosphorylated protein species (DEPs) were identified in leaves of NO- or polyethylene glycol (PEG)-treated wheat seedlings (D), and in control seedlings. From 1396 unique phosphoproteins, 2257 unique phosphorylated peptides and 2416 phosphorylation sites were identified. Of these, 96 DEPs displayed significant changes (≥1.50-fold, p < 0.01). These DEPs are involved in photosynthesis, signal transduction, etc. Furthermore, phosphorylation of several DEPs was upregulated by both D and NO treatments, but downregulated only in NO treatment. These differences affected the chlorophyll A⁻B binding protein, chloroplast post-illumination chlorophyll-fluorescence-increase protein, and SNT7, implying that NO indirectly regulated the absorption and transport of light energy in photosynthesis in response to WD stress. The significant difference of chlorophyll (Chl) content, Chl a fluorescence-transient, photosynthesis index, and trapping and transport of light energy further indicated that exogenous NO under D stress enhanced the primary photosynthesis reaction compared to D treatment. A putative pathway is proposed to elucidate NO regulation of the primary reaction of photosynthesis under WD.

Keywords: 20% PEG-induced water deficiency; Triticum aestivum L.; nitric oxide; phosphoproteomic; primary reaction of photosynthesis.

Figure 1

Change of relative water content (RWC) in leaves of wheat seedlings under normal water conditions and in response to three different treatments at different time points; a total of three independent biological replicates were conducted (n = 10). C, normal water conditions; S, pretreated with 150 µmol/L sodium nitroprusside; D, water deficiency induced by 20% polyethylene glycol (PEG)-6000; S + D, pretreated with 150 µmol/L sodium nitroprusside and water deficiency stress by 20% PEG-6000. Asterisks indicate significant differences among the four treatments at p < 0.05.

Figure 2

Phenotypic (A,B) changes in leaves of wheat seedlings under normal water conditions and in response to three different treatments; a total of three independent biological replicates were used (n = 10). For a detailed description of treatment conditions, please refer to the legend of Figure 1. Different lowercase letters indicate a statistically significant difference at p < 0.05.

Figure 3

Upregulated and downregulated significant phosphorylated peptides among four groups. For a detailed description of treatment conditions, please refer to the legend of Figure 1.

Figure 4

(A) Cluster analysis and (B) functional classification of significant differentially phosphorylated peptides in leaves of winter wheat among four different treatments. The color scale bar at the left of the hierarchical cluster analysis indicates the increased (red) and the decreased (green) peptides. For a detailed description of treatment conditions, please refer to the legend of Figure 1.

Figure 5

(A) Chlorophyll (Chl) content (Chl a, Chl b), (B) net photosynthetic rate (Pn), (C) fluorescence intensity, PI_ABS, P_ET, and (D) RC/ABS in leaves of wheat seedlings under normal water conditions and in response to NO or PEG-induced water deficiency treatments. C, normal water conditions; S, pretreated with 150 µmol/L sodium nitroprusside; D, water deficiency induced by treatment with 20% polyethylene glycol (PEG)-6000; S + D, pretreated with 150 µmol/L sodium nitroprusside and then water deficiency stressed by 20% PEG-6000. O, J, I and P mean that the analysis of the transient-considered fluorescence values at 50 ms (Fo, step O), 2 ms (F2 ms, step J), 30 ms (F30 ms, step I), and maximal level (FM, step P), respectively. For a detailed description of treatment conditions, please refer to the legend of Figure 1. Different lowercase letters in (A,B) indicate a statistically significant difference at p < 0.05.

Figure 6

Schematic representation of NO involved in the regulation of photosynthesis to improve water-deficiency stress resistance. The contribution of NO to photosynthesis primary reaction under water deficit stress flows in the direction indicated by the arrow.