3.1 High Temperature Stress Conditions in Plants

Heat stress has become increasingly prevalent due to global climate change, posing a significant threat to crop growth and agricultural productivity worldwide (Merewitz and Liu 2019; Abd El-Naby et al. 2020). In plants, it is characterized by the accelerated degradation of chlorophyll, reduced photosynthetic efficiency, limited stomatal movement, and restricted water use (Zhang et al. 2017; Wang, Xing, et al. 2022; Wang, Lu, et al. 2022; Manafi et al. 2022). These factors collectively contribute to plant senescence, ultimately compromising nutrient and water use efficiency and leading to increased plant mortality (Liang et al. 2018). Exposure to high temperatures is associated with excessive production of ROS, which causes oxidative damage to essential biomolecules, including nucleic acids, cell membranes, enzymes, and other macromolecules within plant cells (Tiryaki and Keles 2012; Chumikina et al. 2019).

Recent studies have highlighted the protective role of melatonin in mitigating heat stress in various plant species. Shi, Wei, Wang, et al. (2016) observed its anti-aging effect in Arabidopsis, while similar benefits were reported in rice (Liang et al. 2015) and cucumber (Zhang et al. 2013). MET inhibits chlorophyllase and pheophorbide a oxygenase (PaO), enzymes responsible for chlorophyll degradation, helping maintain chlorophyll content and photosynthetic efficiency under stress (Wang et al. 2013).

In tomato seedlings subjected to high-temperature stress, MET significantly accelerated the transcription of key enzymes involved in MET biosynthesis, including ASMT, SNAT, T5H, and TDC (Zhang et al. 2024). Notably, MET enhanced TDC expression by 90.61%, whereas the expression levels of T5H, SNAT, and ASMT increased by factors of 5.65, 3.74, and 7.69, respectively, compared with untreated plants (Jahan et al. 2021). These findings elucidate the regulatory roles of MET in the integration of metabolic pathways that enhance tomato resilience to heat stress.

Overall, enhancing MET levels in crops may represent a viable strategy for mitigating the adverse effects of heat stress, ultimately contributing to improved agricultural productivity under rapidly changing climates. Further investigations into the precise molecular mechanisms underlying these protective effects are essential, particularly in understanding MET's role in regulating antioxidant enzyme activity, heat shock protein expression, stomatal conductance, and hormonal signaling pathways. These insights will help optimize agricultural practices and enhance crop resilience under climate stess.

3.2 Drought Conditions in Plants

Drought stress is a critical environmental challenge that significantly affects the growth, development, and productivity of plants. Physiological and biochemical changes induced by water scarcity lead to a range of detrimental effects, including reduced canopy size, decreased stomatal conductance, impaired photosynthesis, accelerated chlorophyll degradation, leaf senescence, and compromised water and nutrient uptake (Ahmad et al. 2021). Under these conditions, the production of ROS increases, resulting in oxidative stress that can cause widespread damage to cellular structures, including membranes, proteins, and nucleic acids (Wang et al. 2013; Fleta-Soriano et al. 2017; Tiwari et al. 2021).

In response to drought stress, plants initiate a complex regulatory network involving various signaling and sensory molecules such as calcium ions (Ca²⁺), ROS, mitogen-activated protein kinases (MAPKs), and phytohormones like ABA and JA (Alam et al. 2018; Wang, Xing, et al. 2022; Wang, Lu, et al. 2022). This network triggers molecular changes that lead to the expression of stress-responsive genes, including dehydration-responsive element-binding (DREB), WRKY bHLHs, HDs, bZIPs, AP2/EREBPs, MYBs, and NAC TFs, as well as the synthesis of late embryogenesis abundant proteins (LEAs) and abscisic acid (ABA), which are crucial for stress signaling and plant adaptation (Zhang, Zhang, Yang, et al. 2014; Zhang, Cruz De Carvalho, et al. 2014; Zhang, Zhang, Zhao, et al. 2014). Tiwari et al. (2021) highlighted MET as a key signaling molecule that is essential for plants to effectively manage water stress. MET upregulates the expression of crucial genes involved in the AsA-GSH cycle, including dehydroascorbate reductase (DHAR), monodehydroascorbate reductase (MDHAR), and APX, thereby enhancing plant antioxidant capacity (Ye et al. 2016; Khan, Numan, et al. 2020; Khan, Khan, et al. 2020). MET plays a crucial role in maintaining water homeostasis during drought stress (Sharma and Zheng 2019). It influences stomatal closure, which reduces transpiration rates and limits water loss in plants (Dai et al. 2020). By regulating stomatal conductance, MET helps sustain internal water levels, thereby enhancing overall plant resilience to drought conditions (Arnao and Hernández-Ruiz 2019). Furthermore, exogenous MET improved photosynthesis, promoted chlorophyll recovery, stabilized cell structures, and enhanced turgor pressure under water stress conditions (Ding et al. 2018).

Under drought conditions, MET enhanced the chlorophyll content and photosynthetic efficiency, facilitating greater biomass accumulation (Zhang et al. 2020). The regulation of root architecture by MET promotes root elongation and lateral root formation, which are essential for improved water and nutrient absorption (Hossain et al. 2020). Additionally, the overexpression of MzSMT1, a gene involved in MET synthesis, significantly enhances drought resistance in Arabidopsis (Zuo et al. 2014).

MET interacts with other phytohormones and plays a vital role in the enhancement of drought tolerance. It has been shown to modulate ABA levels, thereby promoting stomatal closure and improving water use efficiency (Jahan et al. 2021). The interplay between MET and other signaling molecules such as hydrogen sulfide (H2S) and nitric oxide (NO) further underscores MET's role in plant adaptation to water scarcity (Corpas et al. 2024).

Recent advancements in transcriptomic analyses have revealed that MET treatment during drought stress leads to differential expression of genes associated with stress responses, metabolic processes, and antioxidant defense mechanisms (Daszkowska-Golec et al. 2018; Sharma et al. 2020; Wang, Shao, et al. 2024; Wang, Tanveer, et al. 2024). These genomic changes indicate that MET not only enhances immediate physiological responses but also induces long-term adaptations to drought stress at the molecular level. Exogenous application of MET may alleviate the negative effects of drought stress and improve seedling establishment, elongation, and lateral root formation across various crops, including alfalfa (Niu et al. 2022), tomato (Lee and Back 2019), wheat (Cui et al. 2017), and apples (Wang et al. 2013). Enhancing MET levels in crops is a promising strategy for improving drought tolerance and sustaining agricultural productivity under increasingly arid conditions. Future research should focus on elucidating the precise molecular mechanisms underlying the action of MET to harness its potential for optimizing crop resilience against water scarcity.

3.3 Chilling Stress Conditions in Plants

Chilling stress, defined as exposure to temperatures below the optimal range for plant growth, presents a formidable challenge to agricultural productivity, particularly in crops sensitive to low temperatures. This physiological stress can lead to a spectrum of detrimental effects, including reduced photosynthetic efficiency, impaired respiratory function, destabilization of cellular membranes, and increased oxidative stress due to excessive accumulation of ROS (Pu et al. 2021; Darré et al. 2024). The resulting physiological disruptions not only threaten crop survival but also significantly limit yields, thereby underscoring the urgent need for effective strategies to mitigate chilling-induced damage (Posmyk et al. 2009; Ding et al. 2017).

The efficacy of MET in alleviating oxidative stress under chilling stress conditions has been attributed to its regulatory effects on key antioxidant enzymes, including SOD, CAT, and APX (Gao et al. 2016; Gao et al. 2018; Cao et al. 2019; Ma et al. 2023). Collectively, these enzymes form an intricate antioxidative defense system that plants rely on to counteract oxidative damage caused by chilling conditions.

One of the key functions of MET under chilling stress is stabilizing cellular membranes. Low temperatures often disrupt membrane integrity, increasing permeability and leakage of essential cellular components (Ghorbani et al. 2024). MET contributes to the maintenance of membrane stability by enhancing both fluidity and structural integrity of membranes exposed to low-temperature stress (Jafari et al. 2022). This stabilization is crucial for ensuring the proper functioning of metabolic processes and mitigating the adverse physiological effects of chilling stress in plants (Posmyk et al. 2009; Cao et al. 2019).

In conjunction with its antioxidant and membrane-stabilizing capabilities, MET facilitates the accumulation of osmoprotectants, including proline and soluble sugars. These osmoprotectants are essential for maintaining osmotic balance within plant cells, thereby enabling them to manage both dehydration and osmotic stress that may arise during chilling episodes (Hayat et al. 2022). Proline has demonstrated a strong correlation with enhanced tolerance to chilling stress, functioning to stabilize proteins and cellular structures while also serving as an additional ROS scavenger (Madebo et al. 2021).

Furthermore, MET regulates gene expression in response to cold stress (Bajwa et al. 2014). Recent transcriptomic analyses have indicated that MET treatment upregulates the expression of cold-responsive genes involved in the biosynthesis of osmoprotectants and antioxidants, thereby strengthening the plant's natural defense mechanisms (Kebbeh et al. 2023; Bao et al. 2024). By enhancing the expression of these genes, MET promotes adaptive physiological responses that improve the ability of plants to withstand cold temperatures.

In addition to its protective functions, MET alleviates the negative effects of cold stress on photosynthesis (Su et al. 2024). Chilling stress is known to significantly impair photosynthetic capacity, which in turn reduces energy production and inhibits overall plant growth. However, MET has been shown to upregulate photosynthetic genes and stimulate chlorophyll biosynthesis, thereby enhancing photosynthetic efficiency under low-temperature conditions (Fan et al. 2015; Farouk and Al-Amri 2019; Ding et al. 2022; Ma et al. 2023). This improved photosynthetic performance is crucial because it sustains energy production and supports plant development despite the presence of chilling stress.

Moreover, MET accumulates in plants exposed to extreme cold or freezing conditions, providing critical protection against cellular damage, oxidative stress, and metabolic disruption. For instance, transgenic rice plants with elevated expression of SNAT, a key enzyme in the MET biosynthesis, exhibit greater resistance to freezing stress compared to wild-type counterparts (Kang et al. 2010). This highlights MET's role in modulating plant responses to freezing stress and suggests its potential application in enhancing chilling tolerance through genetic engineering.

By fortifying these defensive mechanisms, MET has emerged as a promising biostimulant to improve cold tolerance in sensitive crop species. Continued investigation of the molecular pathways underlying the protective roles of MET will yield valuable insights into its potential applications as a biostimulant, ultimately enhancing crop resilience in the face of climate-induced chilling stress (Lee and Back 2019; Hayat et al. 2022; Bao et al. 2024).

3.4 Water Logging Stress Conditions in Plants

Waterlogging is a significant abiotic stressor that adversely affects plant physiology by creating low oxygen conditions in the soil. Al Azzawi et al. (2020) reported that waterlogging stress impairs various physiological functions in plants, resulting in reduced fertility and productivity. Around 30%–35% of the world's cultivated land has experienced a decline in fertility and productivity as a result of waterlogging (Kaur et al. 2020). This stress significantly increases the accumulation of ROS by inhibiting gas diffusion and stimulating anaerobic respiration in roots (Wu et al. 2021). Excessive ROS accumulation under anaerobic conditions can cause detrimental effects such as root bending, root rot, and plant wilting (Hossain et al. 2009).

Under hypoxic conditions, increased ethylene production promotes root aerenchyma formation and facilitates plant growth (Sasidharan and Voesenek 2015). In response to waterlogging, plants naturally increase their endogenous levels of MET by upregulating the expression of MET biosynthesis genes including COMT, SNAT, ASMT, T5H, and TDC. The increase in MET levels enhances crop resistance to waterlogging by improving photosynthetic efficiency, activating antioxidant enzymes, and promoting robust root growth (Moustafa-Farag, Mahmoud, et al. 2020).

MET plays a crucial role in activating ROS-scavenging systems in both roots and shoots under waterlogged conditions (Sasidharan and Voesenek 2015; Zeng, Liu, et al. 2022). Exogenous application of MET has been shown to significantly elevate the expression of waterlogged stress-responsive genes in various plant species. For instance, Liu et al. (2023) observed that MET upregulates the expression of OsGH3-2, OsPIN2, and OsPIN1b in rice crops subjected to flooded conditions. Additionally, MET significantly increased soluble sugar content (SSC) and indole-3-acetic acid (IAA) levels while reducing superoxide anion accumulation in the root zone (Zeng, Liu, et al. 2022). This suggests that MET supports normal root growth under hypoxic conditions by activating the auxin signaling pathway and enhancing the antioxidant defense system.

MET not only strengthens antioxidant defenses, but also promotes aerenchyma formation. It enhances internal gas exchange by facilitating oxygen diffusion from the aerial parts of the plants to the submerged roots (Hossain et al. 2009; Gu et al. 2021). This adaptation alleviates hypoxic conditions in waterlogged soils and supports root respiration. Furthermore, MET modulates hormonal signaling pathways crucial for stress responses by interacting with the ethylene and abscisic acid pathways to regulate stomatal closure and conserve water (Moustafa-Farag, Elkelish, et al. 2020; Moustafa-Farag, Mahmoud, et al. 2020).

Overall, MET has emerged as a key regulator of plant resilience to waterlogged stress. Through its antioxidant properties, modulation of physiological responses, and promotion of adaptive root development, MET effectively mitigates the adverse effects of waterlogging in various plant species. Continued research into the specific molecular mechanisms underlying the actions of MET will provide valuable insights for the development of crop varieties with enhanced resilience to water-related stresses, ultimately contributing to sustainable agriculture and food security.

3.5 Salt Stress Conditions in Plants

Salinity is increasingly recognized as a major environmental stressor affecting global food production (Butcher et al. 2016). Its adverse effects worsen each year (El Sabagh et al. 2020). High salinity levels boost the production of ROS. This happens due to increased absorption of Na⁺ and Cl⁻ from the soil. At the same time, salinity reduces the accumulation of Ca²⁺ and K⁺ in plants (Yang and Guo 2018). At elevated salinity concentrations, the toxicity of these ions inhibits plant growth, leading to osmotic and oxidative stress.

Halophytes, or salt-tolerant plants, have evolved various biochemical defense mechanisms to cope with high salinity. These mechanisms include expulsion of excess Na⁺ through specialized antiporters, such as tonoplast NHX1 and plasma membrane SOS1 (Zhang and Shi 2013). They also involve the activation of ROS scavenging pathways and accumulation of osmolytes, including proline. These adaptations help to regulate the physiological responses of halophytes to salt stress (Shafeiee and Ehsanzadeh 2019).

The perception of salt stress by plant roots triggers a cytosolic “Ca²⁺ signature,” which is characterized by an increase in cytosolic calcium concentration ([Ca²⁺]cyt). This signaling pathway is mediated by the calcineurin B-like protein (CBL) family, Ca²⁺-dependent protein kinases (CDPKs), and calmodulin (CaM), which function as Ca²⁺ sensors. These proteins play crucial roles in promoting antioxidant enzyme activity, enhancing proline accumulation, and modifying ion homeostasis under saline conditions (Valivand et al. 2019).

Recent research has demonstrated that exogenous MET mitigates the effects of salt stress in several plant species, including rice, tomato, faba bean, maize, cucumber, and Arabidopsis by modulating various physiological activities (Siddiqui, Alamri, Alsubaie, et al. 2019). Specifically, Liu et al. (2020) reported that MET promoted the expression of 23 calcium-binding protein-related genes and seven CBL/CDPK genes in rice under salt stress conditions. Similar transcriptomic responses have been observed in cassava (Hu et al. 2016), Arabidopsis thaliana (Weeda et al. 2014) and Bermuda grass (Shi, Jiang, et al. 2015).

Under salinity stress conditions, MET treatment upregulates the expression of GhAKT1, GhSOS1, and GhNHX1, thereby promoting cotton seed germination (Shen et al. 2021). These findings underscore that MET plays a positive role in regulating salt stress responses by enhancing calcium-related gene expression, ultimately contributing to improved plant tolerance to saline environments.

The application of MET offers a promising avenue to enhance the resilience of crop species to salt stress. MET can serve as a potential biostimulant to support sustainable agriculture in saline-prone regions by activating various physiological and molecular pathways that regulate ion homeostasis and oxidative stress response. Future studies should focus on elucidating the underlying molecular mechanisms of MET action in different crop species to fully exploit its benefits in agricultural practices.

3.6 Heavy Metal Stress Conditions in Plants

Heavy metal contamination of ecosystems has emerged as a critical threat to crop production. Metals, such as arsenic (As; Farouk and Al-Amri 2019; Samanta et al. 2022), aluminum (Al; Khan, Numan, et al. 2020; Khan, Khan, et al. 2020), copper (Zhang et al. 2022), cadmium (Cd; Tousi et al. 2020), lead (Pb; David et al. 2021), zinc (Zn; Eisalou et al. 2021), nickel (Ni; Jahan et al. 2020), mercury (Hg; Wu et al. 2015), and iron (Fe; Ahammed et al. 2020) pose significant risks to plant health. These metals can accumulate in plant tissues, leading to their entry into the food chain, exacerbating environmental and health concerns for humans? (Wu et al. 2015). Excessive accumulation of heavy metals can disrupt various physiological and biochemical processes within plants, including photosynthesis, nutrient mobilization, and enzyme activity. This disruption results in ionic imbalance, osmotic and oxidative stress, cellular toxicity, and eventual cell and tissue death (Ahmad et al. 2015, 2021).

At sub-millimolar concentrations, heavy metals can induce stomatal closure by blocking ion channels (e.g., La³⁺) and water channels (e.g., Zn²⁺, Pb²⁺, Hg²⁺) and interfering with the movement of anions across the tonoplast (Guo et al. 2023). This interference occurs independently of ABA signaling, which limits gas exchange and exacerbates water stress (Asgher et al. 2014; Khan et al. 2016). Furthermore, heavy metals can regulate ABA-induced stomatal activity by modulating various signaling pathways, establishing a complex relationship between water management and stress mitigation (Gálusová et al. 2020).

Plants have evolved multiple defense strategies in response to heavy metal-induced oxidative stress. These strategies include the activation of antioxidant defense systems, which feature both enzymatic components (such as SOD, APX, and CAT) and non-enzymatic antioxidants (such as GSH, AsA, proline, and carotenoids; Bashri and Prasad 2015). These antioxidants work synergistically to neutralize ROS generated during heavy metal exposure, thereby alleviating oxidative damage (Ahmad et al. 2015).

MET has gained attention because of its role in enhancing plant tolerance to heavy-metal stress. It functions as a potent antioxidant that effectively scavenges ROS, thereby reducing oxidative damage (Moustafa-Farag, Elkelish, et al. 2020; Wiszniewska 2021). By promoting the activity of key antioxidant enzymes, such as SOD, APX, and CAT, MET strengthens the ability of the plant to respond to oxidative stress (Jiang et al. 2016; Li et al. 2016; Zhang et al. 2016). Additionally, MET stimulates the production of non-enzymatic antioxidants, including GSH and ASC, thereby enhancing detoxification mechanisms (Zhang et al. 2016).

MET also facilitates the chelation and detoxification of heavy metals by forming metal–ligand complexes, thereby limiting their bioavailability and toxicity (Khan et al. 2021). This process is often mediated by enhanced synthesis of phytochelatins and metallothioneins, which sequester heavy metals and protect plant cells from toxic effect (Moustafa-Farag, Elkelish, et al. 2020). Moreover, MET has been shown to influence metal uptake and translocation. Research has indicated that MET can modulate the expression of transporter proteins involved in the uptake and sequestration of heavy metals. Additionally, MET enhances the uptake of essential nutrients such as sulfur and iron, which help maintain physiological functions and mitigate stress responses (Zhou et al. 2016; Hasan et al. 2018).

In addition to its role in directly combating oxidative stress and facilitating nutrient uptake, MET affects hormonal signaling pathways associated with stress responses. For example, MET enhances the activity of enzymes crucial for secondary metabolite biosynthesis, such as chalcone synthase (CHS), phenylalanine ammonia-lyase (PAL), and dihydroflavonol reductase (DFR), which are essential for the production of flavonoids and phenolic compounds (Li et al. 2017; Jahan et al. 2020). Under Ni stress, MET has been found to enhance proline accumulation by downregulating proline dehydrogenase (PDH) activity while promoting the activity of pyrroline-5-carboxylate synthase (P5CS; Siddiqui, Alamri, Al-Khaishany, et al. 2019). Proline accumulation further aids in osmotic regulation and metal detoxification.

Moreover, MET improves the activity of key enzymes involved in maintaining photosynthetic efficiency under heavy metal stress, such as ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBisCO) and carbonic anhydrase (CA; Shi, Jiang, et al. 2015). MET plays a vital role in restoring enzymatic activity and promoting chlorophyll synthesis by regulating the biosynthesis of important compounds, such as glycine and succinyl-CoA, which are induced by aminolevulinate synthase (Sarropoulou et al. 2012).

Under heavy metal stress conditions, MET enhances plant resilience by accelerating the expression of genes involved in metal uptake and translocation, ultimately leading to improved growth and development. For example, in the presence of chromium stress, MET-induced regulation of key signaling enzymes enhances seed germination in rice (Li, Ding, et al. 2022).

The multifaceted role of MET in enhancing heavy-metal tolerance in plants is evident through its antioxidant properties, its ability to chelate and detoxify metals, its regulatory effects on metabolic pathways, and the promotion of secondary metabolite production. As a promising biostimulant, MET offers a potential strategy to improve crop resilience and productivity in contaminated agricultural systems. Further research is essential to elucidate the underlying molecular mechanisms of MET in various plant species to effectively manage heavy-metal stress.

4.1 MET and AUX

A well-developed root system is essential for plants to effectively mitigate soil-borne stresses. Research has indicated that exogenous MET significantly promotes the formation of adventitious roots by more than two-fold, and lateral roots by over three-fold, often in coordination with IAA (Koyama et al. 2013). This phenomenon is not limited to A. thaliana transgenic lines, as demonstrated by Zuo et al. (2014), but has also been observed in other species, including pomegranate (Punica granatum; Sarrou et al. 2014) and sweet cherries (Prunus avium; Zuo et al. 2014). Furthermore, Ge et al. (2023) reported that MET-treated tomato (Solanum lycopersicum) seedlings exhibited overexpression of caffeic acid O-methyltransferase 1 (SlCOMT1) and alpha-amylase under salt stress, resulting in an increased accumulation of soluble sugars and enhanced starch breakdown within plant cells.

Recent findings suggest that the synergy between MET and IAA is pivotal for enhancing plant growth and facilitating adaptation to stress conditions. Arnao and Hernández-Ruiz (2018) explored various plant species, including lupin, to assess the similarities between different auxin compounds and MET in terms of their effects on gravitropism, growth, and rooting. They found that MET application altered apical IAA utilization by upregulating several auxin signaling-related genes, such as IAA19 and IAA24, and PIN proteins (PIN1, PIN3, and PIN7; Wang et al. 2016). Notably, MET did not significantly affect root elongation when auxin synthesis was entirely suppressed, highlighting that the effects of MET are contingent on auxin levels (Yang 2017).

The role of MET in stimulating lateral and adventitious root formation has been the subject of extensive research, particularly concerning auxins such as indole-3-acetic acid (IAA), 1-naphthaleneacetic acid, and indole-3-butyric acid (Pelagio-Flores et al. 2012). Consequently, scientists are increasingly considering that MET is a valuable promoter of plant growth, with positive effects observed across various species, including Avena, Hordeum, Triticum, Brassica, Prunus, Punica, and Cucumis, as well as in maize, soybeans, and tomatoes (Arnao and Hernández-Ruiz 2018).

Both MET and IAA are biosynthesized from tryptophan, and L-tryptophan decarboxylase (PSID) catalyzes the conversion of aromatic L-amino acids into tryptamine. Tryptamine is then converted to serotonin by tryptamine 5-hydroxylase (CYP71P1), which serves as a precursor for MET synthesis. In contrast, NADPH oxidoreductase facilitates the production of N-hydroxytryptamine from tryptamine, which is subsequently converted into indole-3-acetaldoxime by N-hydroxytryptamine oxidoreductase, contributing to IAA synthesis.

Recent transcriptomic analyses have shown that MET application upregulates the expression of YUC8, YUC7, YUC4, and YUC3, key genes involved in auxin biosynthesis (Wang et al. 2016). Since tryptophan is a common precursor for both MET and IAA, exogenous MET can suppress IAA biosynthesis, particularly in plants where the zinc finger protein ZAT6 plays a dominant role in IAA signaling (Shi et al. 2018). Notably, mutants of A. thaliana defective in auxin transports-PIN5 (involved in intracellular auxin homeostasis) and WAG1 (regulating auxin-mediated root growth)-as well as TT4 and TT2 (flavonoid-related genes that indirectly affect auxin transport and signaling) exhibit reduced lateral root growth in response to MET. This highlights the crucial role of MET–auxin interaction in root development (Ren et al. 2019).

Similar to IAA, MET accelerates lateral root formation and inhibits primary root development. Liang et al. (2017) observed that MET enhances the expression of multiple genes within the root tip meristem. Although MET concentrations exceeding 10 μM may suppress plant growth, they play a crucial role in maintaining normal physiological functions under stress conditions in species such as Cynodon and A. thaliana under cold stress, Zea mays and Helianthus in saline environments, and Malus in low-quality soil (Bajwa et al. 2014; Shi, Reiter, et al. 2015; Kim et al. 2016; Li et al. 2016).

For example, melatonin-treated tomato plants produced 1.4 to 2.0 times more IAA than the untreated controls (Wen et al. 2016). However, excessive accumulation of MET may lead to a significant reduction in IAA levels, indicating that maintaining MET concentrations below 10 μM is optimal to ensure stress resilience. MET regulates IAA synthesis by upregulating the activity of IAA-amino synthase enzymes.

These studies have shed light on the complex mechanisms by which stress and growth-related signaling pathways interact to regulate gene transcription in plants facing abiotic stress (Pauwels et al. 2010). However, the precise physiological interaction between MET and IAA under stressful conditions remains to be fully elucidated (Pelagio-Flores et al. 2012). Further investigations are crucial to clarify the intricate cross-talk between these important phytohormones and variations in their transcriptional regulation under stress.

4.2 MET and GAs

Previous research focusing on vegetable crops, specifically red cabbage and cucumber, has demonstrated that MET enhances seed germination by increasing the transcription of genes responsible for GA production (Posmyk et al. 2009). Under stress conditions, such as drought or salinity, MET upregulates the expression of key enzymes involved in GA biosynthesis, including GA3ox, GA20ox, and GA2ox (Zhang, Zhang, Yang, et al. 2014; Zhang, Cruz De Carvalho, et al. 2014; Zhang, Zhang, Zhao, et al. 2014). These enzymes are crucial for converting inactive GA into active forms, thereby promoting growth and development under adverse conditions.

Furthermore, studies have reported an increase in the expression of GID, which encodes a soluble GA receptor that interacts with DELLA proteins and GA to form a complex. This interaction prevents DELLA proteins from inhibiting GA signaling, thereby facilitating the growth-promoting effects of GA. Similar responses have also been observed in other plant species, such as rice and apples, under various environmental stress conditions (Mao et al. 2020).

Exogenous MET has been found to trigger defensive biochemical activities and promote growth in fluoride-stressed rice seedlings (IR-64) by upregulating the expression of GA biosynthesis genes, including ASMT (acetylserotonin O-methyltransferase), SNAT, TDC, and GA3ox (Banerjee and Roychoudhury 2019). At the same time, endogenous abscisic acid (ABA) levels decrease because of the downregulation of ABA8ox1 and upregulation of the NCED3 (9-cis-epoxycarotenoid dioxygenase) gene, which is involved in ABA biosynthesis.

Interestingly, under elevated GA and MET conditions, the expression of ABA-dependent stress response genes (OSBZ8, TRAB, and WRKY71) was reduced. This reduction ultimately helps restrict fluoride accumulation in stressed seedlings by reactivating P-H⁺/ATPase expression, which is essential for maintaining ion homeostasis (Banerjee and Roychoudhury 2019).

In addition, research on Linum bicolor has shown that MET treatment enhances seed germination under salinity stress by upregulating the GA biosynthesis genes GA20ox and GA3ox. Moreover, MET influences the expression of ABA metabolism genes by upregulating LbCYP707A1 and LbCYP707A2 involved in ABA degradation and downregulating LbNCED1 and LbNCED3 (ABA biosynthesis genes; Li, Yang, et al. 2019). This dual action of MET not only promotes GA biosynthesis, but also helps modulate ABA levels, thereby fostering better adaptation to saline stress.

These interconnected insights highlight the significant role of MET in mediating GA signaling pathways and ABA-related stress responses, establishing MET as a crucial regulator in enhancing plant resilience and growth under adverse environmental conditions. Future research should explore the molecular mechanisms underlying the interactions between MET and GA, further underscoring its potential as a biostimulant in agricultural practices.

4.3 MET and CKs

Cytokinins (CKs) play a crucial role in delaying senescence in stressed plants by maintaining chlorophyll stability and regulating cell division. While both CKs and MET support chlorophyll retention, research has indicated that MET has a greater capacity than CKs for minimizing chlorophyll loss. Studies of the interactions between MET and CKs have shown that the application of exogenous MET significantly increases CK levels by enhancing the expression of genes responsible for MET biosynthesis, such as TDC and SNAT (Zhang et al. 2017; Ma et al. 2018). However, there appears to be little to no correlation between endogenous MET concentrations and CK levels under unfavorable conditions, suggesting a more complex relationship.

Under stress conditions, MET upregulates genes involved in CK biosynthesis, including LpIPT2 (isopentenyl transferase) and LpOG1, as well as TFs, such as A-ARRs and B-ARRs, which are linked to CK signaling pathways (Samanta and Roychoudhury 2023). This suggests that MET enhances the CK pathway and promotes cellular resilience during stress.

For instance, during heat stress in ryegrass (Lolium perenne L.), the exogenous application of MET has been reported to elevate the levels of both endogenous CKs and MET, while simultaneously reducing ABA content (Zhang et al. 2017). This shift inhibits leaf senescence, as demonstrated by the downregulation of senescence-related genes (Lph36 and LpSAG12.1) and the enhancement of B-ARR signaling, as documented by Zhang et al. (2017). Similarly, Jahan et al. (2021) observed comparable effects in tomato plants, where MET treatment increased levels of GA. The combined increase in MET and GA contributed to a delay in the senescence processes.

Moreover, Ma et al. (2018) found that under drought-stressed conditions, the combined application of CKs and MET positively influenced physiological indicators such as chlorophyll content, photochemical efficiency, and relative water content. This synergy resulted in increased levels of transzeatin riboside and isopentenyl adenine in treated plants, highlighting the beneficial role of MET when used in conjunction with CKs under heat stress conditions.

The interaction between MET and CKs enhances the stress response of plants in a coordinated manner, suggesting the potential for tailored applications of these compounds in agricultural practices. However, maintaining MET concentrations within the optimal range of 0.1–10 μM is essential to maximize the synergistic effects without causing phytotoxicity, as suggested by Arnao and Hernández-Ruiz (2018).

Although CKs do not completely prevent senescence, they effectively delay the onset of stress-related effects by promoting nutrient mobilization and regulating various minerals within the plant. However, the genetic interactions between kinetin and MET require further investigation to fully elucidate their synergistic mechanisms. Gaining insights into these interactions could lead to novel strategies for improving crop resilience and productivity under environmental stress.

4.4 MET and ABA

Kanwar et al. (2018) discovered that ABA and its downstream signaling components, such as ABI3 and ABI5, play a crucial role in mediating antioxidant stress responses as downstream signals of MET. Their findings revealed that MET enhances ABA accumulation under stress conditions, which in turn activates ABI3 and ABI5, leading to the regulation of stress responsive genes and antioxidant enzyme activity. This interplay is particularly significant under conditions of drought, salinity, and oxidative stress. Further supporting this relationship, Li, Guo, et al. (2021) and Li, Jiang, et al. (2021) demonstrated that elevated MET concentrations can inhibit seed germination by modulating ABA signaling pathways, reinforcing the role of MET-ABA crosstalk in plant stress adaptation.

MET influences ABA signaling by altering ABA sensitivity and potentially downregulating enzymes responsible for ABA biosynthesis. One key enzyme in this biosynthetic pathway, 9-cis-epoxycarotenoid dioxygenase (NCED), was notably downregulated following MET treatment. Concurrently, genes involved in ABA catabolism, particularly CYP707 monooxygenases, were upregulated, leading to a marked decrease in ABA levels. This phenomenon has been documented in various crops, including cucumber, cabbage, hickory, and apple (Arnao and Hernández-Ruiz 2021). However, the extent of this response is considerably affected by the specific stress conditions faced by the crops and the concentration of the MET applied.

Additionally, MET plays a regulatory role in the synthesis and activity of ethylene hormones by modulating critical factors, such as AP2a, CNR, NOR, and RIN-like fruit ripening factors (Sun et al. 2020). Notably, in tomato plants, the application of MET activates genes responsible for carotenoid production, thereby enhancing both the nutritional value and aesthetic quality of the fruit (Xu et al. 2019; Verde et al. 2023).

Under heat stress conditions, exogenous application of MET in tomatoes leads to a significant reduction in endogenous abscisic acid (ABA) levels, achieving a decrease of 36.34%. This reduction results from the suppression of ABA biosynthesis and signaling pathways. The greatest upregulation of CYP707A1 (162.33%) and CYP707A2 (160.76%) occurred 6 h after MET treatment compared to that in untreated seedlings, contributing to delayed senescence activities, as noted by Jahan et al. (2021).

In rice, MET promotes improved seed germination under low-temperature stress conditions by enhancing the activities of key enzymes, such as acid phosphatase (ACP), cytochrome-c-oxidase (CCO), succinate dehydrogenase (SDH), and α-amylase (Li, Guo, et al. 2021; Li, Jiang, et al. 2021). Simultaneously, MET inhibited the expression of abi5-1 and abi5-2, which are important regulators of the ABA signaling pathway. Moreover, MET treatment significantly downregulated key ABA biosynthesis genes, including OsAAO, OsNCED1, and OsNCED2, thereby effectively reducing ABA levels in plant cells. Notably, MET upregulates catalase 2 (OsCAT2) and modulates ABA-INSENSITIVE5 (OsABI5), a critical component of ABA-mediated stress signaling. This action helps maintain the balance of ROS during low-temperature stress conditions (Li, Guo, et al. 2021; Li, Jiang, et al. 2021).

In tomatoes, the application of exogenous MET significantly reduces endogenous levels of ABA by 36.34% under heat stress conditions (Jahan et al. 2021). This reduction occurs primarily through the downregulation of ABA biosynthesis and signaling pathways. Remarkably, the highest expression of CYP707A1 (162.33%) and CYP707A2 (160.76%) was recorded six hours after MET treatment compared to untreated plants, indicating a delay in senescence activities within the seedlings (Sun et al. 2020; Verde et al. 2023).

In rice, MET promotes seed germination even under low-temperature stress conditions by enhancing the activities of key enzymes, including ACP, CCO, SDH, and α-amylase (Li, Guo, et al. 2021; Li, Jiang, et al. 2021). This enhancement is achieved through the downregulation of the abi5-1 and abi5-2 genes, which are integral to abscisic acid (ABA) signaling. Concurrently, MET treatment reduces ABA levels by suppressing the expression of genes such as OsAAO, OsNCED1, and OsNCED2. Notably, MET positively regulates catalase 2 (OsCAT2) and significantly upregulates ABA-INSENSITIVE5 (OsABI5), a crucial player in ABA-mediated stress signaling, thereby sustaining the balance of ROS during low-temperature stress (Chen et al. 2021; Jahan et al. 2021; Singh et al. 2022).

Overall, these findings reveal that MET serves as a vital regulator of ABA signaling and metabolism, particularly under varied stress conditions, enhancing plant resilience and promoting growth by fine-tuning the delicate balance of hormonal interactions within the plant system. Further research is needed to fully elucidate the molecular mechanisms underlying the complex relationships between MET, ABA, and other hormones to harness their potential in agricultural applications effectively.

4.5 MET and ETH

MET enhances the synthesis of ethylene in plants by modulating the activity of ripening factors, including AP2a, CNR, NOR, and RIN-like proteins. In tomatoes, MET treatment significantly activates the genes responsible for carotenoid biosynthesis, leading to increased transcription of ACC synthase (ACS) and ACC oxidase (ACO) genes, such as ERF2, EIL1, and EIL2 (Huang et al. 2024). Differential proteomic studies on tomato fruits have indicated that MET prominently influences several proteins associated with ripening pathways (Sun et al. 2020). Similarly, in strawberries, MET has been shown to improve post-harvest quality parameters, enhancing fruit quality indices, increasing levels of bioactive compounds (including phenolic acids, antioxidants, and flavonoids), and extending shelf life (Zahedi et al. 2020).

The biosynthesis of ET from MET involves three enzymatic steps: (i) synthesis of S-adenosyl-l-methionine (S-AdoMet) from MET via S-AdoMet synthetase, (ii) conversion of S-AdoMet to 1-aminocyclopropane-1-carboxylic acid (ACC) by ACC synthase, and (iii) subsequent synthesis of ethylene from ACC, mediated by ACC oxidase (Lin et al. 2009). Furthermore, S-adenosyl-l-methionine (SAM) is implicated in various physiological activities and metabolic reactions (Martínez-López et al. 2008), along with S-AdoMet synthetase, which catalyzes the conversion of MET and ATP to SAM (Fontecave et al. 2004).

Wu and Yang (2019) observed that MET-induced accumulation of ethylene in plant cells mitigated the effects of environmental stress by stabilizing chlorophyll levels and preventing the accumulation of ROS. Additionally, research by (Masood et al. 2012) demonstrated that enhancing ethylene homeostasis in mustard plants exposed to heavy metals, primarily Cd, Zn, and Ni, resulted in reduced photosynthetic inhibition while maintaining elevated levels of glutathione (GSH), as highlighted by Xu et al. (2019).

The regulation of ethylene biosynthesis by MET encompasses the modulation of fruit ripening factors, such as AP2a, CNR, NOR, and RIN-like factors. For instance, in tomatoes, MET treatment activates genes responsible for carotenoid production. In apples, exogenous MET promotes fruit ripening by upregulating the expression of MdACO1 and MdACS1, paralleling the effect of the ethylene precursor ACC (Verde et al. 2023). Similarly, Xu et al. (2019) found that ACC, an ethylene precursor, plays a significant role in the regulation of salt tolerance in grapevines. Furthermore, exogenous MET induces the expression of acetylserotonin methyltransferase (VviASMT), which consequently increases endogenous MET biosynthesis in grapevines (Xu et al. 2019).

Moreover, the synergy between ethylene and MET in regulating salt stress is notable because the application of MET enhances ACC content and overall ET biosynthesis by upregulating ACS1 and MYB108A transcription in both grapevines and tobacco (Lee and Back 2019). MYB108A is intricately associated with the ACS1 promoter, promoting ET production in conjunction with elevated MET concentrations (Sun et al. 2020; Khan et al. 2022). These findings underscore the complex interplay between MET and other signaling molecules, suggesting that this relationship may play a crucial role in regulating ethylene production under various abiotic stress conditions. Further research is warranted to fully elucidate the underlying mechanisms governing these interactions and their potential applications for enhancing crop resilience and quality.

4.6 MET and SA

Salicylic acid (SA) is a crucial component of chromatic acids that plays a significant role in maintaining normal physiological functions during stress conditions. According to Moghaddam et al. (2011) this compound contributes to a range of protective mechanisms, including elevated antioxidant levels, balanced osmotic and redox states, improved membrane integrity, optimal water balance during heat stress, and enhanced photosynthetic activity (Wang et al. 2018).

SA acts as a key regulator of both biotic and abiotic stress responses in plants, as demonstrated by Khan et al. (2021). Their research showed that the levels of SA and MET increased in Arabidopsis plants upon infection with Pseudomonas syringae DC300. Notably, SNAT-knockout mutants exhibited decreased levels of both MET and SA, resulting in heightened susceptibility to pathogen attacks (Lee et al. 2015). Furthermore, the exogenous application of MET has been shown to provide protective effects against the tobacco mosaic virus by promoting the production of both SA and nitric oxide (NO). Zhao et al. (2019) observed that MET treatment significantly reduced viral titers and lower relative levels of viral RNA in infected plants.

Haydari et al. (2019) explored the interaction between exogenous MET and SA in relation to changes in chemical profiles, including antioxidant enzyme activities and essential oil content, in Mentha arvensis L. (var. piperaceous) and Mentha × piperita L. (Mitcham variety) under heat-stress conditions. Their findings indicated that both SA and MET effectively alleviated heat stress by enhancing the activities of SOD and CAT while also increasing essential oil concentrations within plant cells.

The combined application of MET and SA enhances the ability of plants to store essential nutrients, including Fe, N, Zn, P, Cu, and K (Eisalou et al. 2021), while simultaneously limiting the accumulation of sodium ions (Na+) to mitigate the impact of salt stress in wheat crops (Talaat and Shawky 2022). This increased salinity tolerance is facilitated by an enhanced H⁺ pump activity, which leads to decreased electrolyte leakage and reduced SOD content, as well as improved cell membrane stability indices (Mg²⁺/Na⁺, Ca²⁺/Na⁺, and K⁺/Na⁺ ratios) and increased antioxidant enzyme activities. Similarly, (Eisalou et al. 2021) investigated the combined effects of MET and SA on the alleviation of zinc toxicity in safflower seedlings. Their results showed that the synergistic application of MET and SA significantly increased the activities of glycolate enzymes and elevated the levels of ASX and GSH in plants under zinc stress conditions.

These findings underscore the synergistic benefits of MET and SA in enhancing plant resilience and survival, particularly in response to various abiotic stressors. Although the interactions between MET and SA show promising potential in regulating a wide range of stress responses, further comprehensive studies are crucial to fully elucidate the underlying mechanisms and optimize their applications in agriculture. Understanding these interactions could pave the way for innovative strategies for improving plant stress tolerance and productivity under changing environmental conditions.

4.7 MET and JA

Jasmonic acid (JA) and its derivative methyl jasmonate (MeJA) are crucial plant hormones that regulate a broad spectrum of physiological processes, including growth, development, and stress responses. Jasmonates play a key role in enhancing transcriptional activity related to plant defense mechanisms against environmental challenges (Yang et al. 2019; Hu et al. 2020; Li, Guo, et al. 2021; Li, Jiang, et al. 2021).

Recent studies have highlighted the complexity of the interactions between JA and MET. Under abiotic stress conditions, MET treatment has been observed to subtly affect the JA levels (Guo et al. 2021; Ding et al. 2022). Specifically, research indicates that MET reduces JA synthesis and content in Brassica napus subjected to stress. Although the exact dynamics of the MET–JA interaction vary depending on the specific stress conditions, MET appears to modulate JA responses by promoting the production of jasmonate–zinc domain (JAZ) proteins, which inhibit JA signaling pathways (Zhu et al. 2020).

Interestingly, a recent investigation using Arabidopsis revealed that elevated doses of MET (0.1 and 1.0 mM) inhibited primary root growth and reduced the production of JA, CKs, and BRs. In contrast, genes associated with the biosynthesis of GAs, strigolactones, and ethylene were upregulated, indicating MET's nuanced regulatory role of MET in balancing these hormonal pathways (Yang et al. 2019).

Hu et al. (2020) demonstrated that the exogenous application of MET enhances lateral root growth in Cucumis sativus under copper-induced stress by lowering JA levels and altering linolenic acid metabolism. MET significantly reduced the expression of lipoxygenase (LOX)-related genes, which decreased linolenic acid accumulation in plant tissues, thereby mitigating ROS damage. Furthermore, MET influenced the transcriptional expression of cell wall-associated genes, including HD-ZIP, GRAS, AP2/ERF, and BBR/BPS, thereby enhancing plant stress tolerance.

Conversely, when MET is applied through irrigation or foliar spraying to soybean plants experiencing water stress, JA levels increase by 62.6% and 52.5%, respectively (Imran et al. 2021). Similarly, MET increases JA production in rapeseed plants under salt stress by approximately 27% compared to control levels. These findings suggest that MET not only influences the biosynthesis of JA but also regulates its distribution and metabolism in response to various environmental stressors (Tan et al. 2019).

These findings illustrate that MET plays a pivotal role in modulating JA signaling pathways, significantly affecting plant resilience and adaptability under stress conditions. Further studies are required to elucidate the precise molecular mechanisms governing the interactions between MET and JA. Understanding these pathways could lead to improved agricultural practices to enhance crop stress tolerance.

4.8 MET and BRs

Brassinosteroids (BRs) are steroid hormones that play a vital role in the regulation of various physiological processes in plants, including cell division, elongation, and overall growth. They are well known for their protective effects under stress conditions, enhancing plant resilience by initiating MAPK cascades that increase the activity of antioxidant enzymes through the upregulation of TFs. For instance, specific TFs such as MYB, DREB2, WRKY1, and NAC have been shown to be expressed in response to drought stress in oats, whereas OsWRKY30 is similarly activated in rice (Danquah et al. 2014).

Interestingly, high levels of MET have been observed to inhibit the expression of genes involved in BRs and IAA biosynthesis, subsequently slowing root growth (Yang et al. 2021). In Arabidopsis, elevated MET concentrations hinder seedling growth by impeding hypocotyl elongation and altering the transcription of BR biosynthesis genes. This inhibition mimics the effect of brassinazole (BRZ), a known BR biosynthesis inhibitor, suggesting that MET restricts the expression of these BR biosynthesis genes (Xiong et al. 2019).

Studies utilizing RNA interference to knock down SNAT2 in rice have demonstrated that this interference reduces both MET and BRs biosynthesis, leading to the development of dwarf plants deficient in BRs. Notably, the decline in BR levels does not always correspond to diminished MET levels because the reduction in BRs is specific to the biosynthesis pathways of MET. For instance, targeting the expression of COMPT (cinnamate-4-hydroxylase), SNAT2, and T5H results in decreased BR levels, whereas the suppression of SNAT1 and TDC does not affect BR levels (Lee and Back 2016). The response of plants with reduced SNAT2 gene expression resembled that observed with lower BR levels, enhancing the tolerance of plants to abiotic stresses by increasing chlorophyll content and reducing malondialdehyde (MDA) levels (Hwang and Back 2022).

Hwang and Back (2021) examined the interplay between MET biosynthesis and the photoreceptor cryptochrome (CRY). In rice seedlings with CRY1b silenced via RNAi, there was an upregulation of MET biosynthesis genes (COMT, SNAT2, SNAT1, and T5H) owing to the suppression of DWARF and BR biosynthesis genes. The observed decline in BR levels in the CRY1b RNAi lines appears to be predominantly due to the reduction in CRY, rather than a direct decrease in MET, as reduced MET primarily inhibited DWARF4, rather than the overall BR biosynthesis pathway.

Moreover, Hwang and Back (2022) reported that both GAs and BRs promote the biosynthesis of endo-methionine. The application of exogenous BR at a concentration of 0.1 mM resulted in a notable increase in MET levels, rising to 1 mM, which was 40% higher than that of the control. This treatment concurrently augmented the transcription levels of MET biosynthetic genes (TDC1, T5H, and ASMT1) while downregulating the expression of COMT and TDC2. Furthermore, BR treatment modulated the expression of M3H metabolic genes and inhibited the expression of ASDAC metabolic genes. These findings indicate that BRs facilitate MET accumulation by inducing the expression of biosynthetic genes, particularly RAVL1, DWARF4, and D11, rather than suppressing the expression of metabolic genes.

The interaction between MET and BRs represents a complex regulatory network in which both hormones influence each other's biosynthesis and signaling pathways. Understanding these interactions is critical for elucidating the mechanisms underlying plant growth and stress responses, paving the way for improved agricultural practices.