Effect of arbuscular mycorrhizal fungi and phosphorus on drought-induced oxidative stress and 14-3-3 proteins gene expression of Populus cathayana

Abstract

The application of arbuscular mycorrhizal fungi (AM fungi) and phosphorus (P) can improve plant growth under drought stress by upregulating the antioxidant system and osmotic accumulation. The 14-3-3 protein can respond to different abiotic stresses such as low P and drought. The purpose of this experiment was to study the effects of AM fungi (Rhizophagus intraradices) inoculation on reactive oxygen species (ROS) homeostasis, P metabolism, and 14-3-3 gene expression of Populus cathayana at different P levels and drought stress (WW: well-watered and WD: water deficit). Under WD conditions, AM fungi inoculation significantly increased the P content in leaves and roots, but the benefit in roots is limited by the level of P addition, and the roots may have more alkaline phosphatase and phytase under P stress, and these activities in the rhizosphere soil inoculated with AM fungi were stronger. Under WD conditions, the activities of catalase (leaf and root) and peroxidase (root) inoculated with AM fungi were significantly higher than those without inoculation and decreased with P addition. 14-3-3 genes, PcGRF10 and PcGRF11, have a positive correlation with the antioxidant system, osmotic regulation, and P metabolism, which may be more significant after inoculation with AM fungi. Our results provide new insights into the mechanism of ROS homeostasis and P metabolism in mycorrhizal plants under drought stress.

Keywords: 14-3-3 proteins; arbuscular mycorrhizal fungi; drought stress; phosphorus metabolism; reactive oxygen species homeostasis.

FIGURE 1

Growth graph, plant height, and root vitality of Populus cathayana seedlings inoculated with AM fungi under different P addition and water treatment. (A) The growth graphs of NM + WW; (B) the growth graphs of AM + WW; (C) the growth graphs of NM + WD; (D) the growth graphs of AM + WD; (E) plant height; and (F) root vitality. NM + WW, well-watered and non-inoculation treatment; AM + WW, well-watered and AM fungi inoculation treatment; NM + WD, water deficit and non-inoculation treatment; AM + WD, water deficit and AM fungi inoculation treatment. P0, P50, P100, and P150 represent different P content of 0, 50, 100, and 150 mg, respectively. The same lowercase letters following the means indicate non-significant differences in corresponding treatments by Duncan’s multiple range test (P < 0.05). Significance mark of three-factor ANVOA: *P < 0.05, ***P < 0.001, the name of three-factor ANVOA: AM represents inoculation with R. intraradices; drought represents drought treatment; P represents different P addition treatments. The value is expressed as the mean ± standard deviation, n = 3.

FIGURE 2

P content of P. cathayana seedlings and soil inoculated with AM fungi under different P addition and water treatment. (A) Leaf total P content; (B) root total P content; (C) soil total P content; (D) soil available P content. NM + WW, well-watered and non-inoculation treatment; AM + WW, well-watered and AM fungi inoculation treatment; NM + WD, water deficit and non-inoculation treatment; AM + WD, water deficit and AM fungi inoculation treatment. P0, P50, P100, and P150 represent different P content of 0, 50, 100, and 150 mg, respectively. The same lowercase letters following the means indicate non-significant differences in corresponding treatments by Duncan’s multiple range test (P < 0.05). Significance mark of three-factor ANVOA: *P < 0.05, ***P < 0.001, NS is not significant; the name of three-factor ANVOA: AM represents inoculation with R. intraradices; drought represents drought treatment; P represents different P addition treatments. The value is expressed as the mean ± standard deviation, n = 3.

FIGURE 3

Phosphatase content of P. cathayana roots and soil inoculated with AM fungi under different P addition and water treatment. (A) Acid phosphatase activity in the root; (B) alkaline phosphatase activity in the root; (C) acid phosphatase activity in the soil; (D) alkaline phosphatase activity in the soil. NM + WW, well-watered and non-inoculation treatment; AM + WW, well-watered and AM fungi inoculation treatment; NM + WD, water deficit and non-inoculation treatment; AM + WD, water deficit and AM fungi inoculation treatment. P0, P50, P100, and P150 represent different P content of 0, 50, 100, and 150 mg, respectively. The same lowercase letters following the means indicate non-significant differences in corresponding treatments by Duncan’s multiple range test (P < 0.05). Significance mark of three-factor ANVOA: *P < 0.05, **P < 0.01, ***P < 0.001, NS is not significant; the name of three-factor ANVOA: AM represents inoculation with R. intraradices; drought represents drought treatment; P represents different P addition treatments. The value is expressed as the mean ± standard deviation, n = 3.

FIGURE 4

Phytase activity of P. cathayana seedlings and phytase activity and phytic acid content of soil inoculated with AM fungi under different P addition and water treatment. (A) Phytase activity in leaf; (B) phytase activity in root; (C) phytase activity in soil; (D) phytic acid content in the soil. NM + WW, well-watered and non-inoculation treatment; AM + WW, well-watered and AM fungi inoculation treatment; NM + WD, water deficit and non-inoculation treatment; AM + WD, water deficit and AM fungi inoculation treatment. P0, P50, P100, and P150 represent different P content of 0, 50, 100, and 150 mg, respectively. The same lowercase letters following the means indicate non-significant differences in corresponding treatments by Duncan’s multiple range test (P < 0.05). Significance mark of three-factor ANVOA: *P < 0.05, ***P < 0.001, NS is not significant; the name of three-factor ANVOA: AM represents inoculation with R. intraradices; drought represents drought treatment; P represents different P addition treatments. The value is expressed as the mean ± standard deviation, n = 3.

FIGURE 5

Antioxidant enzyme activity in the root of P. cathayana seedlings inoculated with AM fungi under different P addition and water treatment. (A) SOD activity; (B) POD activity; (C) CAT activity. NM + WW, well-watered and non-inoculation treatment; AM + WW, well-watered and AM fungi inoculation treatment; NM + WD, water deficit and non-inoculation treatment; AM + WD, water deficit and AM fungi inoculation treatment. P0, P50, P100, and P150 represent different P content of 0, 50, 100, and 150 mg, respectively. The same lowercase letters following the means indicate non-significant differences in corresponding treatments by Duncan’s multiple range test (P < 0.05). Significance mark of three-factor ANVOA: *P < 0.05, **P < 0.01, ***P < 0.001, NS is not significant; the name of three-factor ANVOA: AM represents inoculation with R. intraradices; drought represents drought treatment; P represents different P addition treatments. The value is expressed as the mean ± standard deviation, n = 3.

FIGURE 6

Oxidative damage material in the root of P. cathayana seedlings inoculated with AM fungi under different P addition and water treatment. (A) H₂O₂ content; (B) O₂^– generation rate; (C) proline content; (D) MDA content. NM + WW, well-watered and non-inoculation treatment; AM + WW, well-watered and AM fungi inoculation treatment; NM + WD, water deficit and non-inoculation treatment; AM + WD, water deficit and AM fungi inoculation treatment. P0, P50, P100, and P150 represent different P content of 0, 50, 100, and 150 mg, respectively. The same lowercase letters following the means indicate non-significant differences in corresponding treatments by Duncan’s multiple range test (P < 0.05). Significance mark of three-factor ANVOA: **P < 0.01, ***P < 0.001, NS is not significant; the name of three-factor ANVOA: AM represents inoculation with R. intraradices; drought represents drought treatment; P represents different P addition treatments. The value is expressed as the mean ± standard deviation, n = 3.

FIGURE 7

The PCA results of physiological indexes of P. cathayana seedlings inoculated with AM fungi under different P concentrations and drought stress. (A) PCA analysis of leaf; (B) PCA analysis of root; (C) PCA analysis of soil. The capital letters and different color markers on the right side of each figure represent A: NM + WW + P0, B: NM + WW + P50, C: NM + WW + P100, D: NM + WW + P150, E: AM + WW + P0, F: AM + WW + P50, G: AM + WW + P100, H: AM + WW + P150, I: NM + WD + P0, J: NM + WD + P50, K: NM + WD + P100, L: NM + WD + P150, M: AM + WD + P0, N: AM + WD + P50, O: AM + WD + P100; P: AM + WD + P150. The different colors ovals in each figure represent pink: NM + WW, yellow: AM + WW, purple: NM + WD, and green: AM + WD.

FIGURE 8

The OPLS-DA results of physiological indexes of P. cathayana seedlings inoculated with AM fungi under different P concentrations and drought stress. (A,D) OPLS-DA analysis of leaf; (B,E) OPLS-DA analysis of root; (C,F) OPLS-DA analysis of soil. The capital letters and different color markers on the right side of each figure represent A: NM + WW + P0, B: NM + WW + P50, C: NM + WW + P100, D: NM + WW + P150, E: AM + WW + P0, F: AM + WW + P50, G: AM + WW + P100, H: AM + WW + P150, I: NM + WD + P0, J: NM + WD + P50, K: NM + WD + P100, L: NM + WD + P150, M: AM + WD + P0, N: AM + WD + P50, O: AM + WD + P100; P: AM + WD + P150. The different colors ovals in each figure represent pink: NM + WW, yellow: AM + WW, purple: NM + WD, and green: AM + WD.

FIGURE 9

The RDA results of 14-3-3 gene expression and physiological indexes of P. cathayana seedlings inoculated with AM fungi under different P concentrations and drought stress. (A,C) RDA analysis of leaf; (B,D) RDA analysis of root. *P < 0.05, **P < 0.01, and ***P < 0.001. The capital letters and different color markers on the right side of each figure represent A: NM + WW + P0, B: NM + WW + P50, C: NM + WW + P100, D: NM + WW + P150, E: AM + WW + P0, F: AM + WW + P50, G: AM + WW + P100, H: AM + WW + P150, I: NM + WD + P0, J: NM + WD + P50, K: NM + WD + P100, L: NM + WD + P150, M: AM + WD + P0, N: AM + WD + P50, O: AM + WD + P100; P: AM + WD + P150. The different colors ovals in each figure represent pink: NM + WW, yellow: AM + WW, purple: NM + WD, and green: AM + WD.