1 Introduction

Since plants are sessile, they regulate their relationships with their environment against the many different environmental stressors they encounter (Abdulkhair and Alghuthaymi 2016; Peck and Mittler 2020). Currently, agricultural areas are becoming unfavorable for plants due to climatic fluctuations or/and anthropogenic interferences that disrupt the hydrological properties of the soil. Salinity and drought are the most common type of stress plants encounter in nature. The high concentration of salts in the soil makes soil water inaccessible to plants due to its low water potential (Sánchez-Romera and Aroca 2020).

Under salinity stress, plants exert adaptive molecular mechanisms to mitigate various morphological and physiological changes (Rahnama et al. 2010; James et al. 2011; Abobatta 2020). Plants respond to salinity at the osmotic and ionic stress tolerance levels. In case of insufficient ionic homeostasis, the increased Na+ content in the cell causes reactive oxygen species (ROS) to accumulate. Accumulated ROS negatively affect cell homeostasis by causing enzyme inactivation and DNA damage. In order to cope with this situation, the osmotic balance is controlled by antioxidant defense systems consisting of both enzymatic and non-enzymatic components. Enzymatic antioxidants form the main line of defense against free radicals in the cell, and in general, the accumulation of compounds is positively correlated against salt stress. At the beginning of these enzymatic antioxidants are Superoxide dismutase (SOD), Catalase (CAT) and Ascorbate peroxidase (APX) enzymes (Shi et al. 2006; Tuteja 2007; Munns and Tester 2008; Isayenkov and Maathuis 2019). SOD has the ability to convert oxygen and hydrogen peroxide. SODs are divided into 3 classes based on their metal cofactors: Cu/Zn-SOD (CSD) (found in cytosol, chloroplast, peroxisome), Mn-SOD (MSD) (found in mitochondria or peroxisome), and Fe-SOD (FSD) (found mostly in mitochondria and chloroplast). The enzyme CAT removes hydrogen peroxide which is reduced by two electrons from oxygen. APX is primarily found in the chloroplast and cytosol and work as the key enzyme of the ascorbate cycle, converting ascorbic acid to dehydroascorbate, eliminating peroxidase (Asada 1992, Willekens et al. 1997; Al-aghabary et al. 2005; Prashanth et al. 2008; Bose et al. 2014; Ighodaro and Akinloye 2018).

Silicon (Si) has vital roles in plants’ life as a macro element. This element is the most abundant element in various compound forms on earth after oxygen, and constitutes 25–30% of the earth’s crust. Plantstake Si from the soil as silicic acid [Si(OH)4], an uncharged monomeric molecule, under less than 9 pH (Shi et al. 2013; Luyckx et al. 2017; Tripathi et al. 2017; Saitoh et al. 2021). Although the beneficial effects of Si are minimal under optimal conditions, the rise in stress factors (abiotic and/or biotic) increases the importance of Si for both agricultural and horticultural activities. It is stated that the harvest quality of various crops is increased by fertilization using silicon. However, the mechanisms of Si-mediated stress responses are still not fully elucidated (Savvas et al. 2009; Ashraf et al. 2010; Bakhat et al. 2018; Siam et al. 2018).

Many stages of development and responses to environmental cues are regulated directly or indirectly by epigenetic mechanisms (Sudan et al. 2018). DNA methylation is the most studied type of epigenetic modification in the cell. It occurs via the transfer of the methyl group from S-adenosyl methionine (SAMe) to cytosines by methyltransferase enzymes after replication. Demethylation of DNA can be passive and/or active in plants. Passive DNA demethylation occurs when methyltransferases are inactive during the cell cycle following DNA replication, leaving the newly synthesized strand unmethylated. Active DNA demethylation occurs when DNA glycosylases, which are normally associated with DNA repair, recognize 5-methylcytosine and remove the methyl group from DNA. DNA demethylation can activate expressed alleles of some imprinted genes (Gehring et al. 2009; Zhu 2009). DNA methylation in plants occurs in two types; symmetrical CG, CHG, and asymmetrical CHH (where H represents A, T, or C) (Kong et al. 2018). With the completion of A. thaliana genome sequencing in 2000, it was revealed that there are three main enzymes responsible for the methylation of cytosines in plants. These three methylation enzymes are: METHYLTRANSFERASE 1 (MET1), plant-specific CHROMOMETHYLASE 3 (CMT3) and DOMAINS REARRANGED METHYLASE 2 (DRM2). Plant-specific Pol IV and Pol V, developed from Polymerase II, are involved in RNA-mediated DNA methylation (RdDM) (Chinnusamy and Zhu 2009; Law and Jacobsen 2010). Also, four DNA demethylase genes have been identified in Arabidopsis, namely DEMETER (AtDME), SILENCING REPRESSOR (AtROS1), DEMETER LIKE 2 (AtDML2) and DEMETER LIKE 3 (AtDML3) (Chan et al. 2005). ROS1 acts as a transcriptional gene silencing repressor by catalyzing the removal of 5-mC and actively demethylating target DNA without the need for replication via a base excision repair pathway (Bharti et al. 2015).

The molecular mechanism of silicon has not yet been elucidated. Although the impact of changes in methylation level on gene regulation is well known, there are extremely limited studies on methylation level changes related to Si metabolism. In this study, the effect of methylation on silicon metabolism was examined by studying two methylation and one demethylation mutants of the Arabidopsis plant. To examine the salt stress mitigating effects of Si, combined salt (NaCl) and silicon [Si(OH)4] treatment was applied to Arabidopsis thaliana methylation mutants (met1-7, drm2-2, and ros1-4). Then, osmolyte accumulation and ion leakage analyzes were performed within the scope of testing physiological changes in mutant plants. The expression of CSD2, CAT3 and APX1 genes, which encode enzymes involved in antioxidant pathways, was examined. Additionally, methylation changes of the mutants used in this study after Si and/or NaCl treatment were analyzed at the global DNA methylation level.

2 Materials and Methods

2.1 Plant Material and Definition of Mutants

The seeds used in this study are mutants of the genes MET1, DRM2 and ROS1 all of which are Columbia-0 ecotypes. Previously described A. thaliana met1-7, drm2-2 and ros1-4 mutants were used as the experimental group while Col-0 (Columbia-0) plants were used as the control group [Col-0 seeds obtained by Dr. Ralf Stracke [The Center for Biotechnology (CeBiTec), Bielefeld University]. The characteristics of the homozygous seeds, NASC codes, Salk numbers are given in Table 1.

Table 1 Arabidopsis epigenetic mutants
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2.2 Seed Germination, Growth Conditions and Stress Treatment

All seeds were surface sterilized and placed on to Murashige and Skoog medium (MS) (Murashige and Skoog 1962) for in vitro culturing. Col-0, met1-7, drm2-2 and ros1-4 seeds were incubated for 7 days to germinate under fluorescent light in a plant growth chamber [16 h light / 8 h dark conditions, 105 µmol m− 2s− 1 (Sanyo, MLR-352 H)] at 25 °C.

Arabidopsis plants were transferred to the medium prepared including 100 mM NaCl (Merck, 1.06404), 1 mM Si (Sigma Aldrich, 306,363) and 100 mM NaCl + 1 mM Si for stress application and incubated for 14 days. After stress treatment, plants were used directly for physiological analysis, the rest was stored at -80 °C to use in molecular analysis. For Si application, silicic acid, which is the form in which plants can uptake the easiest from the soil, was used (Luyckx et al. 2017; Tripathi et al. 2017).

2.3 Physiological Analysis

The osmolyte accumulation was measured using the semi-micro osmometer (Knauer K-7400) as stated by Dourado et al. (2019), and ion leakage in the cell membrane wasmeasured using a conductivity meter (HORIBA B-137) as described by Shi et al. (2006).

2.4 DNA Isolation and Global DNA Methylation (5-mC) Level Analysis

DNA isolation from ∼ 100 mg (is equal to 2 plants for Col-0 and 4 or 5 plants for mutants) of plants harvested after treatment was performed with a GeneJET Plant Genomic DNA Purification Kit (Thermo Scientific, K0792) according to the manufacturer’s protocol. Then, the DNA samples were checked for their integrity, purity and quantity. Their integrities were analyzed by agarose gel electrophoresis and their purities as well as quantities were checked by NanoDrop 2000 (Thermo Scientific). The DNA samples were stored at -20 °C to be used in the methylation experiment. Global methylation changes (%) in genomic DNA were detected with the MethylFlash Global DNA Methylation (5-mC) ELISA Easy Kit (EpiGentek, p-1030) using an indirect ELISA method which is based on antigen-antibody reactions, according to the manufacturer recommended instructions using 100 ng of gDNA for each sample.

2.5 RNA Isolation and qPCR Analysis

After treatment, 100 mg (is equal to 2 plants for Col-0 and 4 or 5 plants for mutants) plant samples were collected from all experimental groups for isolation of RNA with Hibrizol (Hibrigen, Türkiye). Their integrity was checked with agarose gel electrophoresis (1%). Their purity was checked using a NanoDrop 2000 (Thermo Scientific) and their quantity was measured. After quantitative and qualitative controls, cDNA synthesis was performed with the High-Capacity cDNA Reverse Transcription Kit (Thermo Scientific, 4,368,814).

The gene expression profiles of CSD2, CAT3 and APX1 genes were evaluated in met1-7, drm2-2 and ros1-4 epigenetic mutants and Col-0. The primer sequences of the selected genes were designed using the Primer3 program (http://primer3.ut.ee/). Their sequences and product lengths are given in Table 2. qPCR experiments were performed with the LightCycler Nano instrument (Roche, Basel, Switzerland) using 2X SYBR Green qPCR Mix (Hibrigen, Türkiye). The Actin 8 gene was chosen as the endogenous control for normalization of qPCR results. The cycling conditions were set as hold (50 °C, 60 s), denaturation (94 °C, 30 s.), annealing (59 °C for Actin 8, CAT3 and APX1; 68 °C for CSD2 30 s.), extension (72 °C, 30 s.), and hold (40 °C, 30 s.), respectively. The Ct values were evaluated relative to the Actin 8 gene using the 2−ΔΔCt method (Livak and Schmittgen 2001).

Table 2 The primer sequences used in gene expression analysis
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2.6 Statistical Analysis

Physiological experiments with Col-0, met1-7, ros1-4 and drm2-2 plants, global DNA methylation level and gene expression analysis were performed in three biological and two technical repetitions. The data obtained in the experiments were statistically analyzed using GraphPad Prism® 9.0.0 software using a one-way ANOVA with the post-hoc Tukey’s test. P values less than 0.001 are given three asterisks, and P values less than 0.0001 are given four asterisks (ns P > 0,05, * P ≤ 0,05, ** P ≤ 0,01, *** P ≤ 0,001, **** P ≤ 0,0001).

3 Results

3.1 Physiological Effects of Stress Application

The analyzes to determine osmolyte accumulation and ion leakage were performed to evaluate the physiological effects of Si, NaCl and the combined Si and NaCl treatments on epigenetic mutants. For this purpose, osmolyte accumulation and ion leakage were evaluated between the experimental groups.

It is observed that NaCl application in Col-0 plants causes a significant increase compared to control and combined applications of Si and NaCl, while Si treatment does not cause a significant change at the end of (Fig. 1). A significant increase was observed in the met1-7 mutant only between the control and NaCl treatment. In the drm2-2 mutant, Si application caused a significant decrease compared to the control group, while NaCl application caused a significant increase of osmolytes. It was determined that combined Si and NaCl application significantly reduced the amount of osmolytes compared to NaCl application. This showed us that Si has a mitigating effect on NaCl stress in the drm2-2 mutant. In ros1-4 mutants, as compared to the control, the application of Si and NaCl resulted in a significant increase in osmolytes. When the osmolyte accumulation data were examined in general, it was observed that NaCl application caused an increase in the amount of osmolytes in all epigenetic mutants, similar to the Col-0 plant.

Fig. 1
figure 1

The osmolyte accumulation (mOsmol/kg) of the Col-0, met1-7, drm2-2, and ros1-4 plants following treatments is shown graphically. The statistical significance levels of the data are indicated by varying numbers of asterisks

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According to the ion leakage analysis results shown in Fig. 2, it was observed that Si applied to Col-0 and drm2-2 mutants did not cause ion leakage, but in the met1-7 mutant, Si had a slightly increasing effect on ion leakage. While an increase in ion leakage was observed after NaCl application in Col-0 and ros1-4 mutants, a mitigating effect of Si was observed when Si and NaCl was applied simultenously. Salt stress significantly increased ion leakage in all experimental groups except the drm2-2 mutant.

Fig. 2
figure 2

The percentage of ion leakage in Col-0, met1-7, drm2-2, and ros1-4 under different treatments is shown graphically. The statistical significance levels of the data are indicated by varying numbers of asterisks

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3.2 Global DNA Methylation

Global DNA methylation analysis showed that Si application caused significant degree of hypermethylation in all the experimental groups except the ros1-4 mutants (Fig. 3). On the other hand, while NaCl application caused hypomethylation in all groups (more severe in epigenetic mutants), the combined applications of Si and NaCl eliminated the hypomethylation effect of NaCl.

Fig. 3
figure 3

The percentage (%) changes in global DNA methylation of Col-0, met1-7, drm2-2, and ros1-4 under different treatments is shown graphically. The statistical significance levels of the data are indicated by varying numbers of asterisks

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3.3 Changes in Gene Expression Profiles

It was observed that when Si was applied to the Col-0 plant alone, CSD2 gene expression which is one of the initial steps of the antioxidant process activated following stress in plants, did not change, but increased when Si and NaCl applied simultaneously. In the met1-7 mutant, unlike the Col-0 plant, it was seen that all the applied treatments upregulated the gene expression and the maximum level was reached in the combined Si and NaCl application. It might be suggested that by activating the antioxidant system in the met1-7 mutant, silicon exerts its mitigating effect on NaCl stress. A similar situation was seen in the drm2-2 mutant as in Col-0. While there was no significant increase in gene expression when Si was applied alone, it was observed that NaCl application caused a significant increase, however, the maximum level was reached in combined Si and NaCl application. In the ros1-4 mutant, similar to drm2-2, Si application did not cause a significant change in CSD2 gene expression, while NaCl and combined Si and NaCl application appeared to upregulate gene expression. As a result, the increase in the amount of gene expression of CSD2 in all experimental groups confirms that Si has a mitigating effect on salt stress (Fig. 4).

Fig. 4
figure 4

Different experimental treatements exerted differential changes in the expression of CSD2 in (A) Col-0, (B) met1-7, (C) drm2-2 and (D) ros1-4 is shown graphically. The statistical significance levels of the data are indicated by varying numbers of asterisks

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For the CAT3 gene, which plays a role in another step of the antioxidant mechanism, it was observed that combined Si and NaCl application caused a significant increase in Col-0 plants compared to Si and NaCl applications. In the met1-7 mutant, it was observed that NaCl application significantly increased CAT3 gene expression compared to both Si and combined Si and NaCl applications. It was observed that Si and NaCl applications induced a significant increase in CAT3 expression in drm2-2 mutant compared to the control group. Si and NaCl combined tretament significantly down-regulated CAT3 gene expression compared to both Si and NaCl applied separately. In the ros1-4 mutant, similar to drm2-2, NaCl application caused a significant increase compared to the control group. It was observed that the level of CAT3 gene expression caused by Si application was significantly lower than the level of gene expression in NaCl and the combined Si and NaCl applications (Fig. 5).

Fig. 5
figure 5

The variations in CAT3 gene expression levels in (A) Col-0, (B) met1-7, (C) drm2-2 and (D) ros1-4 grown under different experimental conditions is shown graphically. The statistical significance levels of the data are indicated by varying numbers of asterisks

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Considering that Si application along with NaCl stress showed the alleviating effect of Si via upregulating CAT3 gene expression in the Col-0 plant.

For the APX1 gene, which is involved in another step of the antioxidant mechanism, similar to CAT3, Si application in Col-0 plants caused a significant decrease in gene expression level compared to control and NaCl application. It was also observed that the combined Si and NaCl application caused a significant decrease compared to the control group. In the ros1-4 mutant, it was seen that NaCl application caused a significant increase compared to the control group and combined Si and NaCl application. APX1 gene level showed a similar pattern in all experimental groups (Fig. 6).

Fig. 6
figure 6

The changes in the gene expression of APX1 in (A) Col-0, (B) met1-7, (C) drm2-2 and (D) ros1-4 grown under different experimental treatments is shown graphically. The statistical significance levels of the data are indicated by varying numbers of asterisks

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4 Discussion

Silicon, the second most abundant element in soil, has mostly been researched in terms of its role in plant absorption and its potential protective mechanisms against various stressors (Epstein 1994; Thorne et al. 2020; Celayir et al. 2023). It was demonstrated that silicon exerts a mitigating effect on salt stress, however the underlying molecular mechanisms remain partially understood (Liang et al. 1996; Goto et al. 2003; Tahir et al. 2006, 2012; Parveen and Ashraf 2010; Bae et al. 2012; Yin et al. 2013). In plants, salt stress causes ionic stress by disrupting the ionic balance of Na+, Cl−, K+ and Ca2+ together with osmotic stress. An excessive amount of Na+ ions entering the cells is transported to other leaves, tissues and organs where it changes the metabolisms in various ways. This interference causes physiological and molecular effects in plants. Studies have shown that silicon reduces the effects of salt stress by increasing K+ uptake against excessive Na+ intake to the plant (Liang et al. 1996; Zuccarini 2008; Yang and Guo 2018; Liu et al. 2019). However, although the effect of salt stress on epigenetic mechanisms is mostly studied, the effects of Si on epigenetic mutants exposed to salt stress have not been studied. In this study, we tried to understand the physiological and molecular effects of Si on methylation mutants under salt stress. We selected three epigenetic mutants with ecotypes Col-0 as plant materials; these mutants were met1-7, drm2-2 and ros1-4 with loss of function in MET1, DRM2 and ROS1 enzymes by T-DNA insert, respectively.

Osmolytes majorly contribute to maintaining cellular osmotic adjustment through cell turgidity, protect internal cell components and reduce ionic toxicity. In most of the plants, osmolyte accumulation is a common response after exposure to salinity stress. They increase their osmolytes like proline and glycine betaine in order to ameliorate the devastating effects of stress (Singh et al. 2022). An increase in the amount of osmolytes may indicate that stress response mechanisms are actively working (Kosar et al. 2019). In previous studies, it was stated that Si application increased the amount of osmolyte and the osmotic potential (Yin et al. 2013; Farouk et al. 2020). Abbas et al. (2015) examined the alleviating effect of Si on salt stress in okra. Spraying Si on leaves in salinity conditions significantly increased osmolytes and caused an increase in osmotic adaptation capacity and antioxidant activity. In line with our research, Celayir et al. (2023) observed in their study on the effects of Si on UV stress that the application of Si led to a higher concentration of osmolytes compared to the plants that were not treated with Si. However, when UV-B and Si were applied together, the amount of osmolyte decreased, whereas in our results the amount of osmolyte increased. Although the application of Si silicon alone in the drm2-2 mutant causes a decrease in the amount of osmolyte, it confirms that Si applied together with NaCl in Col-0 and drm2-2 reveals its protective effect under stress. The fact that changes in the amount of osmolyte occur at different rates in the mutantsshows that the activation of biomolecules effective in the absorption of silicon and salt might be affected by regional methylation differences.

After stress exposure, ion homeostasis as well as cell membrane permeability of the cell change. This ion flux is often interrelated with accumulation of ROS in the cell. In previous studies using various plants, it has been reported that Si application causes improvement in ion leakage caused by salt stress (Liang et al. 1996; Agarie et al. 1998; Reezi et al. 2009; Kim et al. 2014). Zhu et al. (2004) applied 1 mM Si, 50 mM NaCl and combination treatment to varieties of cucumber and obtained similar data with our results in the ion leakage analysis they performed on the 5th and 10th days of culture. Abdelaal et al. (2020) stated that when Si was added to the medium, ion leakage decreased in sweet pepper plants, while ion leakage increased at different salt stress conditions. Compared to our findings, similar results were observed in the Col-0 plant and the demethylation mutant ros1-4. On the other hand, it was found that Si application under salt stress in methylation mutants met1-7 and drm2-2 further increases ion leakage. This can be interpreted as the loss of methylation prevents Si from affecting membrane permeability, regardless of which regions are methylated.

DNA methylation in plants plays a crucial role in the regulation of many epigenetic phenomena. These include transcriptional silencing of transposons and transgenes, defense against stress conditions, silencing of genes that control seed formation and leaf morphology at flowering time (Zangi et al. 2020). Tobiasz-Salach et al. (2023) examined the physiological, biochemical, and epigenetic effects of Si under salinity stress. In order to identify alterations in DNA methylation in maize plants treated with Si under salt stress, they employed the methylation-sensitive amplified polymorphism (MSAP). According to their findings, plants respond to salt stress by altering their DNA methylation level, and the effects of NaCl together Si application were dose-dependent. They came to the conclusion that these modifications might point to pathways for salt-stressed plant adaptation. In addition, they discovered that applying Si topically can lessen the detrimental effects of salt in the soil, which is in line with previous studies. In another study, the goal of Stadnik et al. (2022) was to assess how silicon foliar treatment affected the oats’ (Avena sativa L.) physiological and epigenetic response to salt stress. Similar to the study conducted by Tobiasz-Salach et al. (2023), the impact of Si administration during salt stress on the DNA methylation level in oat plants was examined using MSAP analysis. Similarly, their research showed that the oat plants’ ability to withstand salt was enhanced by the exogenous application of silicon. This study examined both the physiological and epigenetic responses of plants to the experimental stimuli used. The lowering level of methylation is influenced by the administration of Si under saline conditions. Moreover, Celayir et al. (2023) reported that 1 mM Si application to Arabidopsis Col-0 plant caused hypermethylation, while Yeşildirek et al. (2023) reported that 100 mM NaCl application to met1-7 mutant caused hypomethylation. Also, Yeşildirek et al. (2023) stated in their study that the met1-7 mutant might have supported the genes involved in the RdDM pathway to cope with the DNA methylation deficiency in the CG regions. The fact that the drm2-2 mutant which was in this study gave parallel results with the met1-7 mutant among the applied NaCl condition suggests that a similar situation may also be in question for the drm2-2 mutant. It was observed that the hypermethylation state caused by salt stress was reversed in all epigenetic mutants in the presence of salt stress and Si.

The enzymatic antioxidants that work the most as a defense system against ROS accumulation are SOD, CAT and APX1 enzymes (Maqsood et al. 2023). In previous studies on various plants, it has been shown that the application of Si together with salt causes an increase in antioxidant enzyme activities, leading to an improvement in ROS accumulation (Zhu et al. 2004; Alzahrani et al. 2018; Ahmad et al. 2019; Alamri et al. 2020; Hurtado et al. 2020; Hernández-Salinas et al. 2022). In parallel with our results, Ma et al. (2016) reported thatTaCAT andTaSOD genes expression levels in wheat were increased in high levels when Si was applied to drought-stressed plants than to Si alone was treated. In our study, while no significant change was observed in theAPX gene in the Col-0 plant under salt stress, it was observed that theAPX gene expression increased under salt stress in ros1-4 mutant. It is thought that the gene expression enhancing effect of the epigenetic mutants used in the study was observed due to methylation defects in the gene regions associated with CAT3 expression.

Global methylation analysis showed that combined Si and NaCl application eliminated the hypomethylation caused by NaCl. CSD2 gene expression increased more in Si-NaCl combined application than in NaCl stress alone, regardless of the difference in epigenetic regulation differences. While CAT3 gene expression did not differ in control under NaCl stress, it increased in all epigenetic mutants in the study. In Si-NaCl combined application, this increase was suppressed to a certain extent. The APX gene, on the other hand, increased in NaCl application in mutants, and this increase was suppressed when Si and NaCl were applied together.

Plant responses to stress factors are greatly influenced by DNA methylation, one of the main epigenetic regulation mechanisms. Changes in DNA methylation also enable plants to withstand adverse environmental conditions and develop ‘epigenetic memory’ for the next stress situation. With this study, it was observed that Si, which has previously proven its ameliorating effect against the physiological and morphological negative effects of salt stress, can provide the same effect in terms of global DNA methylation. This suggests that RNA-directed DNA methylation (RdDM) pathway-based regulation may play an important role in the Si-Stress-Plant interaction when methylation and demethylation enzymes do not work properly.

5 Conclusion

One of the primary epigenetic control systems, DNA methylation, has a significant impact on how plants respond to stressors. Findings reveal that applying Si to Arabidopsis methylation mutants during salt stress increases the resistance of plants to salty conditions, depending on the methylation level at the epigenetic level. This study sheds light on the mitigating properties of Si, specifically the methylation that occurs in response to separate and simultaneous exposure to Si and NaCl. This gives us proof that the Si-Stress-Plant relationship may be significantly regulated by the RNA-directed DNA methylation (RdDM) pathway. In conclusion, it was found that treating Arabidopsis epigenetic mutants with Si during salt stress could improve the plants’ ability to withstand salt on epigenetic level.Moreover, our research provides an epigenetic perspective, providing an additional contribution to the mechanisms by which Si promotes plant resilience in stressful environments.