Hydrogen Sulfide Alleviates Manganese Stress in Arabidopsis

Hydrogen sulfide (H₂S) has been shown to participate in various stress responses in plants, including drought, salinity, extreme temperatures, osmotic stress, and heavy metal stress. Manganese (Mn), as a necessary nutrient for plant growth, plays an important role in photosynthesis, growth, development, and enzymatic activation of plants. However, excessive Mn²⁺ in the soil can critically affect plant growth, particularly in acidic soil. In this study, the model plant Arabidopsis thaliana was used to explore the mechanism of H₂S participation and alleviation of Mn stress. First, using wild-type Arabidopsis with excessive Mn²⁺ treatment, the following factors were increased: H₂S content, the main H₂S synthetase L-cysteine desulfhydrase enzyme (AtLCD) activity, and the expression level of the AtLCD gene. Further, using the wild-type, AtLCD deletion mutant (lcd) and overexpression lines (OE5 and OE32) as materials, the phenotype of Arabidopsis seedlings was observed by exogenous application of hydrogen sulfide donor sodium hydrosulfide (NaHS) and scavenger hypotaurine (HT) under excessive Mn²⁺ treatment. The results showed that NaHS can significantly alleviate the stress caused by Mn²⁺, whereas HT aggravates this stress. The lcd mutant is more sensitive to Mn stress than the wild type, and the overexpression lines are more resistant. Moreover, the mechanism of H₂S alleviating Mn stress was determined. The Mn²⁺ content and the expression of the Mn transporter gene in the mutant were significantly higher than those of the wild-type and overexpression lines. The accumulation of reactive oxygen species was significantly reduced in NaHS-treated Arabidopsis seedlings and AtLCD overexpression lines, and the activities of various antioxidant enzymes (SOD, POD, CAT, APX) also significantly increased. In summary, H₂S is involved in the response of Arabidopsis to Mn stress and may alleviate the inhibition of Mn stress on Arabidopsis seedling growth by reducing Mn²⁺ content, reducing reactive oxygen species content, and enhancing antioxidant enzyme activity. This study provides an important basis for further study of plant resistance to heavy metal stress.     

Single Amino Acid Exchange in ACTIN2 Confers Increased Tolerance to Oxidative Stress in Arabidopsis der1–3 Mutant

Single-point mutation in the ACTIN2 gene of the der1–3 mutant revealed that ACTIN2 is an essential actin isovariant required for root hair tip growth, and leads to shorter, thinner and more randomly oriented actin filaments in comparison to the wild-type C24 genotype. The actin cytoskeleton has been linked to plant defense against oxidative stress, but it is not clear how altered structural organization and dynamics of actin filaments may help plants to cope with oxidative stress. In this study, we characterized root growth, plant biomass, actin organization and antioxidant activity of the der1–3 mutant under oxidative stress induced by paraquat and H₂O₂. Under these conditions, plant growth was better in the der1–3 mutant, while the actin cytoskeleton in the der1–3 carrying pro35S::GFP:FABD2 construct showed a lower bundling rate and higher dynamicity. Biochemical analyses documented a lower degree of lipid peroxidation, and an elevated capacity to decompose superoxide and hydrogen peroxide. These results support the view that the der1–3 mutant is more resistant to oxidative stress. We propose that alterations in the actin cytoskeleton, increased sensitivity of ACTIN to reducing agent dithiothreitol (DTT), along with the increased capacity to decompose reactive oxygen species encourage the enhanced tolerance of this mutant against oxidative stress.     

Melatonin Modulates Plant Tolerance to Heavy Metal Stress: Morphological Responses to Molecular Mechanisms

Heavy metal toxicity is one of the most devastating abiotic stresses. Heavy metals cause serious damage to plant growth and productivity, which is a major problem for sustainable agriculture. It adversely affects plant molecular physiology and biochemistry by generating osmotic stress, ionic imbalance, oxidative stress, membrane disorganization, cellular toxicity, and metabolic homeostasis. To improve and stimulate plant tolerance to heavy metal stress, the application of biostimulants can be an effective approach without threatening the ecosystem. Melatonin (N-acetyl-5-methoxytryptamine), a biostimulator, plant growth regulator, and antioxidant, promotes plant tolerance to heavy metal stress by improving redox and nutrient homeostasis, osmotic balance, and primary and secondary metabolism. It is important to perceive the complete and detailed regulatory mechanisms of exogenous and endogenous melatonin-mediated heavy metal-toxicity mitigation in plants to identify potential research gaps that should be addressed in the future. This review provides a novel insight to understand the multifunctional role of melatonin in reducing heavy metal stress and the underlying molecular mechanisms.     

A Novel DUF569 Gene Is a Positive Regulator of the Drought Stress Response in Arabidopsis

In the last two decades, global environmental change has increased abiotic stress on plants and severely affected crops. For example, drought stress is a serious abiotic stress that rapidly and substantially alters the morphological, physiological, and molecular responses of plants. In Arabidopsis, several drought-responsive genes have been identified; however, the underlying molecular mechanism of drought tolerance in plants remains largely unclear. Here, we report that the “domain of unknown function” novel gene DUF569 (AT1G69890) positively regulates drought stress in Arabidopsis. The Arabidopsis loss-of-function mutant atduf569 showed significant sensitivity to drought stress, i.e., severe wilting at the rosette-leaf stage after water was withheld for 3 days. Importantly, the mutant plant did not recover after rewatering, unlike wild-type (WT) plants. In addition, atduf569 plants showed significantly lower abscisic acid accumulation under optimal and drought-stress conditions, as well as significantly higher electrolyte leakage when compared with WT Col-0 plants. Spectrophotometric analyses also indicated a significantly lower accumulation of polyphenols, flavonoids, carotenoids, and chlorophylls in atduf569 mutant plants. Overall, our results suggest that novel DUF569 is a positive regulator of the response to drought in Arabidopsis.     

Cloning of the Lycopene β-cyclase Gene in Nicotiana tabacum and Its Overexpression Confers Salt and Drought Tolerance

Carotenoids are important pigments in plants that play crucial roles in plant growth and in plant responses to environmental stress. Lycopene β cyclase (β-LCY) functions at the branch point of the carotenoid biosynthesis pathway, catalyzing the cyclization of lycopene. Here, a β-LCY gene from Nicotiana tabacum, designated as Ntβ-LCY1, was cloned and functionally characterized. Robust expression of Ntβ-LCY1 was found in leaves, and Ntβ-LCY1 expression was obviously induced by salt, drought, and exogenous abscisic acid treatments. Strong accumulation of carotenoids and expression of carotenoid biosynthesis genes resulted from Ntβ-LCY1 overexpression. Additionally, compared to wild-type plants, transgenic plants with overexpression showed enhanced tolerance to salt and drought stress with higher abscisic acid levels and lower levels of malondialdehyde and reactive oxygen species. Conversely, transgenic RNA interference plants had a clear albino phenotype in leaves, and some plants did not survive beyond the early developmental stages. The suppression of Ntβ-LCY1 expression led to lower expression levels of genes in the carotenoid biosynthesis pathway and to reduced accumulation of carotenoids, chlorophyll, and abscisic acid. These results indicate that Ntβ-LCY1 is not only a likely cyclization enzyme involved in carotenoid accumulation but also confers salt and drought stress tolerance in Nicotiana tabacum.     

ZmCIPK21, A Maize CBL-Interacting Kinase,Enhances Salt Stress Tolerance in Arabidopsis thaliana

Salt stress represents an increasing threat to crop growth and yield in saline soil. In this study, we identified a maize calcineurin B-like protein-interacting protein kinase (CIPK), ZmCIPK21, which was primarily localized in the cytoplasm and the nucleus of cells and displayed enhanced expression under salt stress. Over-expression of ZmCIPK21 in wild-type Arabidopsis plants increased their tolerance to salt, as supported by the longer root lengths and improved growth. The downstream stress-response genes, including dehydration-responsive element-binding (DREB) genes were also activated in transgenic plants over-expressing ZmCIPK21. In addition, introduction of the transgenic ZmCIPK21 gene into the Arabidopsis mutant cipk1-2 rescued the salt-sensitive phenotype under high salt stress. Measurement of Na⁺ and K⁺ content in transgenic plants showed that over-expression of ZmCIPK21 decreased accumulation of Na⁺ and allowed retention of relatively high levels of K⁺, thereby enhancing plant tolerance to salt conditions.     

Polyamine Oxidase Triggers H2O2-Mediated Spermidine Improved Oxidative Stress Tolerance of Tomato Seedlings Subjected to Saline-Alkaline Stress

Saline-alkaline stress is one of several major abiotic stresses in crop production. Exogenous spermidine (Spd) can effectively increase tomato saline-alkaline stress resistance by relieving membrane lipid peroxidation damage. However, the mechanism through which exogenous Spd pre-treatment triggers the tomato antioxidant system to resist saline-alkaline stress remains unclear. Whether H₂O₂ and polyamine oxidase (PAO) are involved in Spd-induced tomato saline-alkaline stress tolerance needs to be determined. Here, we investigated the role of PAO and H₂O₂ in exogenous Spd-induced tolerance of tomato to saline-alkaline stress. Results showed that Spd application increased the expression and activities of superoxide dismutase (SOD), catalase (CAT), ascorbate peroxidase (APX), glutathione reductase (GR), and the ratio of reduced ascorbate (AsA) and glutathione (GSH) contents under saline-alkaline stress condition. Exogenous Spd treatment triggered endogenous H₂O₂ levels, SlPAO4 gene expression, as well as PAO activity under normal conditions. Inhibiting endogenous PAO activity by 1,8-diaminooctane (1,8-DO, an inhibitor of polyamine oxidase) significantly reduced H₂O₂ levels in the later stage. Moreover, inhibiting endogenous PAO or silencing the SlPAO4 gene increased the peroxidation damage of tomato leaves under saline-alkaline stress. These findings indicated that exogenous Spd treatment stimulated SlPAO4 gene expression and increased PAO activity, which mediated the elevation of H₂O₂ level under normal conditions. Consequently, the downstream antioxidant system was activated to eliminate excessive ROS accumulation and relieve membrane lipid peroxidation damage and growth inhibition under saline-alkaline stress. In conclusion, PAO triggered H₂O₂-mediated Spd-induced increase in the tolerance of tomato to saline-alkaline stress.     

Flavonone 3-hydroxylase Relieves Bacterial Leaf Blight Stress in Rice via Overaccumulation of Antioxidant Flavonoids and Induction of Defense Genes and Hormones

Efficient accumulation of flavonoids is important for increased tolerance to biotic stress. Although several plant defense mechanisms are known, the roles of many pathways, proteins, and secondary metabolites in stress tolerance are unknown. We generated a flavanone 3-hydroxylase (F3H) overexpressor rice line and inoculated Xanthomonas Oryzae pv. oryzae and compared the control and wildtype inoculated plants. In addition to promoting plant growth and developmental maintenance, the overexpression of F3H increased the accumulation of flavonoids and increased tolerance to bacterial leaf blight (BLB) stress. Moreover, leaf lesion length was higher in the infected wildtype plants compared with infected transgenics. Kaempferol and quercetin, which scavenge reactive oxygen species, overaccumulated in transgenic lines compared with wildtypes in response to pathogenic infection, detected by scanning electron microscopy and spectrophotometry. The induction of F3H altered the antioxidant system and reduced the levels of glutathione peroxidase activity and malondialdehyde (MDA) contents in the transgenic lines compared with the wildtypes. Downstream gene regulation analysis showed that the expression of F3H increased the regulation of flavonol synthase (FLS), dihydroflavonol 4-reductase (DFR), and slender rice mutant (SLR1) during BLB stress. The analysis of SA and JA signaling revealed an antagonistic interaction between both hormones and that F3H induction significantly promoted SA and inhibited JA accumulation in the transgenic lines. SA-dependent nonexpressor pathogenesis-related (NPR1) and Xa1 showed significant upregulation in the infected transgenic lines compared with the infected control and wildtype lines. Thus, the overexpression of F3H was essential for increasing BLB stress tolerance.     

Heterologous Expression of the Melatonin-Related Gene HIOMT Improves Salt Tolerance in Malus domestica

Melatonin, a widely known indoleamine molecule that mediates various animal and plant physiological processes, is formed from N-acetyl serotonin via N-acetylserotonin methyltransferase (ASMT). ASMT is an enzyme that catalyzes melatonin synthesis in plants in the rate-determining step and is homologous to hydroxyindole-O-methyltransferase (HIOMT) melatonin synthase in animals. To date, little is known about the effect of HIOMT on salinity in apple plants. Here, we explored the melatonin physiological function in the salinity condition response by heterologous expressing the homologous human HIOMT gene in apple plants. We discovered that the expression of melatonin-related gene (MdASMT) in apple plants was induced by salinity. Most notably, compared with the wild type, three transgenic lines indicated higher melatonin levels, and the heterologous expression of HIOMT enhanced the expression of melatonin synthesis genes. The transgenic lines showed reduced salt damage symptoms, lower relative electrolyte leakage, and less total chlorophyll loss from leaves under salt stress. Meanwhile, through enhanced activity of antioxidant enzymes, transgenic lines decreased the reactive oxygen species accumulation, downregulated the expression of the abscisic acid synthesis gene (MdNCED3), accordingly reducing the accumulation of abscisic acid under salt stress. Both mechanisms regulated morphological changes in the stomata synergistically, thereby mitigating damage to the plants’ photosynthetic ability. In addition, transgenic plants also effectively stabilized their ion balance, raised the expression of salt stress–related genes, as well as alleviated osmotic stress through changes in amino acid metabolism. In summary, heterologous expression of HIOMT improved the adaptation of apple leaves to salt stress, primarily by increasing melatonin concentration, maintaining a high photosynthetic capacity, reducing reactive oxygen species accumulation, and maintaining normal ion homeostasis.     

Crosstalk between Melatonin and Reactive Oxygen Species in Plant Abiotic Stress Responses: An Update

Melatonin acts as a multifunctional molecule that takes part in various physiological processes, especially in the protection against abiotic stresses, such as salinity, drought, heat, cold, heavy metals, etc. These stresses typically elicit reactive oxygen species (ROS) accumulation. Excessive ROS induce oxidative stress and decrease crop growth and productivity. Significant advances in melatonin initiate a complex antioxidant system that modulates ROS homeostasis in plants. Numerous evidences further reveal that melatonin often cooperates with other signaling molecules, such as ROS, nitric oxide (NO), and hydrogen sulfide (H₂S). The interaction among melatonin, NO, H₂S, and ROS orchestrates the responses to abiotic stresses via signaling networks, thus conferring the plant tolerance. In this review, we summarize the roles of melatonin in establishing redox homeostasis through the antioxidant system and the current progress of complex interactions among melatonin, NO, H₂S, and ROS in higher plant responses to abiotic stresses. We further highlight the vital role of respiratory burst oxidase homologs (RBOHs) during these processes. The complicated integration that occurs between ROS and melatonin in plants is also discussed.     

Melatonin Protects Tobacco Suspension Cells against Pb-Induced Mitochondrial Dysfunction

Recent studies have shown that melatonin is an important molecule in plant physiology. It seems that the most important is that melatonin effectively eliminates oxidative stress (direct and indirect antioxidant) and switches on different defence strategies (preventive and interventive actions) during environmental stresses. In the presented report, exogenous melatonin potential to protect Nicotiana tabacum L. line Bright Yellow 2 (BY-2) exposed to lead against death was examined. Analyses of cell proliferation and viability, the level of intracellular calcium, changes in mitochondrial membrane potential (ΔΨm) as well as possible translocation of cytochrome c from mitochondria to cytosol and subsequent caspase-like proteolytic activity were conducted. Our results indicate that pretreatment BY-2 with melatonin protected tobacco cells against mitochondrial dysfunction and caspase-like activation caused by lead. The findings suggest the possible role of this indoleamine in the molecular mechanism of mitochondria, safeguarding against potential collapse and cytochrome c release. Thus, it seems that applied melatonin acted as an effective factor, promoting survival and increasing plant tolerance to lead.