Impact of Drought on Soluble Sugars and Free Proline Content in Selected Arabidopsis Mutants

Abstract

Water shortage is an increasing problem affecting crop yield. Accumulation of compatible osmolytes is a typical plant response to overcome water stress. Sucrose synthase 1 (SUS1), and glucan, water dikinase 2 (GWD2) and δ¹-pyrroline-5-carboxylate synthetase 1 (P5CS1) are members of small protein families whose role in the response of Arabidopsis thaliana plants to mild osmotic stress has been studied in this work. Comparative analysis between wild-type and single loss-of-function T-DNA plants at increasing times following exposure to drought showed no differences in the content of water-insoluble carbohydrate (i.e., transitory starch and cell wall carbohydrates) and in the total amount of amino acids. On the contrary, water-soluble sugars and proline contents were significantly reduced compared to wild-type plants regardless of the metabolic pathway affected by the mutation. The present results contribute to assigning a physiological role to GWD2, the least studied member of the GWD family; strengthening the involvement of SUS1 in the response to osmotic stress; showing a greater contribution of soluble sugars than proline in osmotic adjustment of Arabidopsis in response to drought. Finally, an interaction between proline and soluble sugars emerged, albeit its nature remains speculative and further investigations will be required for complete comprehension.

Keywords: drought stress; metabolic adjustment; proline; soluble sugars; water-insoluble carbohydrates.

Figure 1

Relative water content (WC) under osmotic stress. Relative WC was calculated on whole rosettes cut from 20–30 plants for each genotype at each experimental point. Plants were collected at 12 h light under control condition (CTR) and at 0.5, 4.5 and 6.5 days after treatment (DAT). The fresh weights (FWs) were determined weighting plants immediately after the excision, while the dry weights (DWs) were determined after drying (24 h incubation at 80 °C). Water content was calculated as the difference between the FW and the DW of each plant and expressed as a percentage of the FW. Data are reported in boxplots, the central line represents the median and error bars the highest and the lowest data of the set. The t-test (wild-type vs. T-DNA lines) was used for statistics: * p < 0.01; ** p < 0.001. p-values are reported in Table S1.

Figure 2

Lipid peroxidation under osmotic stress. The degree of stress induced by mannitol treatment was assessed by measuring lipid peroxidation in Arabidopsis rosettes collected at 12 h light under control condition (CTR) and at 0.5, 4.5 and 6.5 DAT. On average, 5 independent biological samples were analyzed for each experimental point. Data are reported in boxplots, the central line represents the median and error bars show the highest and the lowest data of the set. The t-test (wild-type vs. T-DNA lines) was used for statistics: * p < 0.01; ** p < 0.001. p-values are reported in Table S2.

Figure 3

Carbohydrate pools rearrangements under water deprivation. Quantification of leaf starch (a), total water-soluble sugars (b), glucose pool (c) and fructose pool (d) in Arabidopsis plants grown under control condition (CTR) and exposed to mannitol treatment at 0.5, 4.5 and 6.5 DAT. Plants were collected at 12 h light. On average, 5 independent biological samples were analyzed for each experimental point. Data are reported in boxplots, the central line represents the median and error bars show the highest and the lowest data of the set. The t-test (wild-type vs. T-DNA lines) was used for statistics: * p < 0.01; ** p < 0.001. p-values are reported in Table S3.

Figure 4

Cell wall (CW) carbohydrates. Quantification of insoluble non-starch carbohydrates from Arabidopsis plants grown under control condition (CTR) and exposed to mannitol treatment at 0.5, 4.5 and 6.5 DAT. Arabidopsis rosettes were collected at 12 h light. On average, 4 independent biological samples were analyzed for each genotype at each experimental point. Data are reported in boxplots, the central line represents the median and error bars show the highest and the lowest data of the set. The t-test (wild-type vs. T-DNA lines) was used for statistics. p-values are reported in Table S4.

Figure 5

Amino acids and proline accumulation. Quantification of free amino acids (a) and proline (b) in Arabidopsis leaves collected at 12 h light under control condition (CTR) and at 0.5, 4.5 and 6.5 DAT. On average, 3 independent biological samples were analyzed for each genotype at each experimental point. Data are reported in boxplots, the central line represents the median and error bars show the highest and the lowest data of the data set. The t-test (wild-type vs. T-DNA lines) was used for statistics: * p < 0.01; ** p < 0.001. p-values are reported in Table S5.