Physiological and transcriptomic analysis of antioxidant mechanisms in sweet sorghum seedling leaves in response to single and combined drought and salinity stress

The effects of drought, salinity, and combined stress on ROS and ROS metabolic physiology and transcriptomics in sweet sorghum seedling leaves were evaluated. The results showed that drought stress had little effect on photosynthesis, while the SOD activity, CAT activity, and the expression of their related genes were elevated in leaves, but no excessive accumulation of O2-, H2O2, or -OH was observed. Under salinity stress, photosynthesis was inhibited, the O2-, H2O2 and -OH contents increased significantly, and the SOD, POD, CAT activities and the expression of their related genes in leaves were elevated. Under combined stress, photosynthesis was significantly inhibited, the highest accumulation of O2-, H2O2 and -OH contents occurred, and the SOD and POD activities and the expression of related genes in leaves were significantly increased, but the CAT was significantly decreased. These results collectively indicate that oxidative damage to sweet sorghum seedling leaves was higher with combined stress than with either drought or salinity stress alone. Under combined stress the SOD and POD activities were increased, but the CAT activity in the AsA-GSH cycle was severely reduced, demonstrating that antioxidant mechanisms in seedlings did not play a normal protective role, leaving the plants severely damaged by oxidative stress. Abbreviations: AsA: Ascorbic acid; APX: Ascorbate peroxidase; CAT: Catalase; Chl a: Chlorophyll a; Chl b: Chlorophyll b; Ci: Intercellular CO2 concentration; DHAR: Dehydroascorbate reductase; DEGs: Differentially expressed genes; DW: Dry weight; ETR: Photosynthetic electron transport rate; FW: Fresh weight; Fv/Fm: Original light energy conversion efficiency; GPX: Glutathione peroxidase; GR: Glutathione reductase; Gs: Stomatal conductance; GSH: Oxidized glutathione; H2O2: Hydrogen peroxide; MDA: Malondialdehyde; MDHAR: Monodehydroascorbate reductase; O2-: Superoxide anion; PCA: Principal component analysis; PCD: Programed cell death; PEG-6000: Polyethylene glycol 6000; POD: Peroxidase; Pn: Photosynthetic rate; Pro: Proline; qRT–PCR: Quantitative real-time PCR; RNA-seq: RNA Sequencing; ROS: Reactive oxygen species; RWC: Relative water content; RMP: Relative membrane permeability; SOD: Superoxide dismutase; SW: Saturation weight; Tr: Transpiration rate; ΦPSII: Actual photochemical efficiency; -OH: Hydroxyl radical

本研究评估了干旱、盐胁迫及其复合胁迫对甜高粱幼苗叶片活性氧（Reactive oxygen species, ROS）代谢生理及转录组学的影响。结果表明：干旱胁迫对幼苗叶片光合作用影响微弱，叶片中超氧化物歧化酶（Superoxide dismutase, SOD）、过氧化氢酶（Catalase, CAT）活性及其相关基因表达量均有所升高，但未观测到超氧阴离子（Superoxide anion, O₂⁻）、过氧化氢（Hydrogen peroxide, H₂O₂）与羟基自由基（Hydroxyl radical, ·OH）的过量积累。盐胁迫下，叶片光合作用受到抑制，O₂⁻、H₂O₂及·OH含量显著升高，SOD、过氧化物酶（Peroxidase, POD）、CAT活性及其相关基因表达量均有所提升。复合胁迫下，叶片光合作用受到显著抑制，O₂⁻、H₂O₂与·OH含量达到最高积累水平，SOD、POD活性及其相关基因表达量显著升高，但CAT活性显著降低。

上述结果表明，复合胁迫对甜高粱幼苗叶片造成的氧化损伤显著高于单一干旱或盐胁迫。复合胁迫下，尽管SOD与POD活性有所升高，但抗坏血酸-谷胱甘肽（Ascorbic acid-Glutathione, AsA-GSH）循环中的CAT活性严重受损，说明幼苗的抗氧化防御机制未能发挥正常保护作用，致使植株遭受严重的氧化胁迫损伤。

缩写说明：
AsA：抗坏血酸（Ascorbic acid）；
APX：抗坏血酸过氧化物酶（Ascorbate peroxidase）；
CAT：过氧化氢酶（Catalase）；
Chl a：叶绿素a（Chlorophyll a）；
Chl b：叶绿素b（Chlorophyll b）；
Ci：胞间CO₂浓度（Intercellular CO₂ concentration）；
DHAR：脱氢抗坏血酸还原酶（Dehydroascorbate reductase）；
DEGs：差异表达基因（Differentially expressed genes）；
DW：干重（Dry weight）；
ETR：光合电子传递速率（Photosynthetic electron transport rate）；
FW：鲜重（Fresh weight）；
Fv/Fm：原初光能转化效率（Original light energy conversion efficiency）；
GPX：谷胱甘肽过氧化物酶（Glutathione peroxidase）；
GR：谷胱甘肽还原酶（Glutathione reductase）；
Gs：气孔导度（Stomatal conductance）；
GSH：氧化型谷胱甘肽（Oxidized glutathione）；
H₂O₂：过氧化氢（Hydrogen peroxide）；
MDA：丙二醛（Malondialdehyde）；
MDHAR：单脱氢抗坏血酸还原酶（Monodehydroascorbate reductase）；
O₂⁻：超氧阴离子（Superoxide anion）；
PCA：主成分分析（Principal component analysis）；
PCD：程序性细胞死亡（Programmed cell death）；
PEG-6000：聚乙二醇6000（Polyethylene glycol 6000）；
POD：过氧化物酶（Peroxidase）；
Pn：光合速率（Photosynthetic rate）；
Pro：脯氨酸（Proline）；
qRT–PCR：实时定量聚合酶链反应（Quantitative real-time PCR）；
RNA-seq：RNA测序（RNA Sequencing）；
ROS：活性氧（Reactive oxygen species）；
RWC：相对含水量（Relative water content）；
RMP：相对膜透性（Relative membrane permeability）；
SOD：超氧化物歧化酶（Superoxide dismutase）；
SW：饱和重量（Saturation weight）；
Tr：蒸腾速率（Transpiration rate）；
ΦPSII：实际光化学效率（Actual photochemical efficiency）；
·OH：羟基自由基（Hydroxyl radical）