Introduction

In recent years, increasing research attention has been paid to salt stress that can affect the growth and development of plants, especially in the stage of seed germination [1, 2]. The response of plants to salt stress is complex, which is achieved through a variety of physiological and biochemical reactions and molecular mechanisms, and also involves the differential expression of stress-related genes in the corresponding pathways [3,4,5].

Soluble sugars as osmoprotectants help in the alleviation of the negative effects of salt on plants. Rosa et al. [6] suggested that there was a significant increase in the levels of glucose, sucrose, and fructose in plants under salt stress, which played a vital role in carbon storage, osmotic protection, and free radical scavenging. Nemati et al. [7] demonstrated that exogenous glucose can limit the accumulation of Na+ and stimulate the uptake of K+, thereby maintaining ion homeostasis under salt stress.

Trehalose was first isolated from ryegrass by Wiggers in the late nineteenth century and was later found in bacteria, algae, yeast, lower plants, insects, and other invertebrates [8]. The amount of trehalose present in plants is extremely low, and excessive accumulation of trehalose can affect the metabolism of normal carbohydrates in plants and even inhibit their growth [9,10,11]. Trehalose produces a marked effect primarily in the form of signaling molecules under stress conditions [12]. The metabolic pathway of trehalose in plants is recognized as the Ots A–Ots B pathway, in which under the action of trehalose phosphate synthase, uridine diphosphate glucose and glucose-6-phosphate form trehalose-6-phosphate and uridine diphosphate and then generate trehalose and inorganic phosphates under the catalysis of trehalose phosphatase, as shown in Fig. 1 [13, 14]. Garcia et al. [15] reported in 1997 that when rice was exposed to salt stress, the addition of low concentrations of trehalose (1–10 mmol/L) could protect its root integrity, protect root cells from abnormal cell division induced by severe salt stress, and excrete excessive Na+ from cells by protecting ion pumps. Yang et al. [16] demonstrated that trehalose can increase the salt tolerance of plants, while exogenous trehalose may act as an inducer of biological processes involved in the salt stress response of plant.

Fig. 1
figure 1

Metabolic pathway of trehalose in plants

Full size image

Sugars not only participate in the metabolism of carbon and energy in cells but also play a key role in plant growth and stress resistance as a signal molecule [17]. The presence of an interaction has been suggested between sugars and plant hormones in metabolic pathways and stress response [18, 19]. Cheng et al. [20] found that exogenous glucose enhanced the expression of genes related to the biosynthesis of abscisic acid (ABA) and ultimately increased the ABA content in plants.

However, the effects of exogenous trehalose on sugar metabolism and the interaction between trehalose and ABA under salt stress are still unclear. In previous studies, we used transcriptome sequencing to screen the pathways and the related genes involved in protein metabolism and carbohydrate metabolism pathways in tomato [21]. In response to salt stress, the protein metabolism pathway was significantly enriched in seven pathways, with 17 key genes being differentially expressed, and the carbohydrate metabolism pathway was significantly enriched in six pathways, with 19 key genes being differentially expressed. Based on transcriptome sequencing results, this study analyzed the metabolic changes in sugar and ABA in tomato seedlings treated with trehalose and salt and determined the differential expression profiles of key genes involved in sugar metabolism. The purpose of this study was to investigate the effects of trehalose on salt stress and the interaction between sugar metabolism and trehalose on regulating the response to salt stress in tomato, which could provide a theoretical basis for improving crop yield, salt tolerance of plants, and the utilization rate of saline–alkali land in agricultural research.

Materials and Methods

Plant Material and Treatment

After soaking for 4–6 h at room temperature, tomato (cv. Ailsa Craig) seeds were sterilized with 75% ethanol and 10% NaClO, cultured at 25 °C for 8 d, and then cultured in Hoagland’s nutrient solution for 4 weeks with illumination for 16 h/d at 25 °C. Trehalose and salt were applied to tomato seedlings with consistent growth, and each treatment was performed for 7 d as follows: (T1) control: 0 mmol/L Tre + 0 mmol/L NaCl; (T2) 0 mmol/L Tre + 200 mmol/L NaCl; (T3) 1 mmol/L Tre + 200 mmol/L NaCl; (T4) 2 mmol/L Tre + 200 mmol/L NaCl; (T5) 3 mmol/L Tre + 200 mmol/L NaCl; (T6) 5 mmol/L Tre + 200 mmol/L NaCl; and (T7) 10 mmol/L Tre + 200 mmol/L NaCl. Salt concentration was selected based on our previous research [21]. The formulation of Hoagland’s nutrient solution was as follows: pH 6.0, 945 mg/L Ca(NO3)2, 607 mg/L KNO3, 115 mg/L (NH4)3PO4, 493 mg/L MgSO4, 2.5 mL/L ferric salt solution, and 5 mL/L trace element.

Determination of Growth Indicators and Related Physiological Indicators

Plant height, fresh mass, and dry mass of tomato seedlings were measured, and the relative water content was calculated as follows:

$$\begin{aligned} {\text{Relative }}\;{\text{water}}\;{\text{content}} &= \left[ {\left( {{\text{fresh}}\;{\text{mass}} - {\text{dry }}\;{\text{mass}}} \right)/{\text{fresh}}\;{\text{mass}}} \right]\\&\quad \times 100\% \end{aligned}$$
(1)

The chlorophyll content of tomato leaves was measured by the acetone method as previously described in Ref. [22] with slight modification. The activity of peroxidase (POD) was determined by the guaiacol method, and the activity of superoxide dismutase (SOD) was determined by the photochemical reaction method of nitroblue tetrazolium according to previously described methods, with slight modification [23, 24]. Catalase kit was used to determine the activity of CAT enzyme according to the instructions (Solarbio, Beijing, China). The level of proline was determined as described in Ref. [25].

For the determination of malondialdehyde (MDA) levels, 1.0 g seedlings was added to 10 mL of 10% trichloroacetic acid solution, ground on ice, and then centrifuged at 5000 g. Then, 1 mL supernatant was aspirated and 3 mL of 0.6% thiobarbituric acid solution was added and heated in boiling water for 15 min. After cooling the liquid rapidly, absorbance values at 450, 532, and 600 nm were measured. The MDA level was determined according to the following formula:

$$c\left( {\upmu{\text{mol}}/{\text{L}}} \right) = 6.45 \times \left( {{\text{OD}}_{532} - {\text{OD}}_{600} } \right) - 0.56 \times {\text{OD}}_{450}$$
(2)

The optimum trehalose treatment concentration was selected using the technique for order preference by similarity to ideal solution (TOPSIS) analysis.

Determination of Starch and Soluble Sugar Contents Under Optimal Trehalose Concentration

Tomato seedlings (cultivated as mentioned earlier) with consistent growth were selected for four treatments (0 mmol/L Tre + 0 mmol/L NaCl; 0 mmol/L Tre + 200 mmol/L NaCl; 2 mmol/L Tre + 0 mmol/L NaCl; and 2 mmol/L Tre + 200 mmol/L NaCl), and each treatment was processed separately for 0, 1, 3, 5, and 7 d. Both roots and leaves of tomato were selected for determination.

Starch and soluble sugar contents were evaluated based on a previously reported protocol, with some modifications [25].

For the determination of trehalose content, 2 g tomato seedlings were ground into powder in a mortar with liquid nitrogen and homogenized by adding 2 mL of 80% ethanol and 1% polyvinylpolypyrrolidone (PVPP). The supernatant was concentrated in 200 mL distilled water and filtered by a microporous membrane (0.45 μm). Trehalose content was determined using the trehalose assay kit (Seebio, China).

Determination of the Genes Expression Related to Sugar Metabolism Under Optimal Trehalose Concentration

Total RNA in tomato leaves and roots was extracted using the Column Plant RNAOUT kit (Tiandz, Beijing, China), and cDNA was synthesized following the instruction of the FastQuant RT kit (Tiangen Biotech, Beijing, China). CDS sequences of internal reference gene and target genes were searched in the NCBI database. The Primer 5.0 software was used to design specific primer sequences given in Table 1. PCR amplification and quantitative real-time PCR (qRT-PCR) were performed using S1000 thermal cycler (BIO-RAD, USA) and LightCycler96 (Roche, Basel, Switzerland), respectively. β-Tublin was selected as the internal reference gene [26].

Table 1 Primer sequences of genes related to sugar metabolism
Full size table

Determination of the Genes Expression Related to Sugar Transporter Under Optimal Trehalose Concentration

The expression of genes related to sugar transporter was measured as mentioned earlier. The primer sequences are given in Table 2.

Table 2 Primer sequences of genes related to sugar transporter
Full size table

Determination of ABA Content and the Expression of Metabolism-Related Genes Under Optimal Trehalose Concentration

After being ground with liquid nitrogen, 0.1 g of tomato seedlings was added with 0.8 mL of 10% methanol and 1% acetic acid, and then extracted at 4 °C for 16 h. After centrifuged at 10000g for 15 min, the supernatant was aspirated and filtered through a 0.45-μm microporous membrane. The ABA content was determined using a Plant ABA ELISA test kit (Jianglai Biotechnology, Shanghai, China).

The expression of metabolism-related genes was determined as mentioned earlier. The primer sequences are given in Table 3.

Table 3 Primer sequences of genes related to ABA metabolism
Full size table

Statistical Analysis

All experiments were repeated three times. Significance of differences (p < 0.05) between the mean values was determined by analysis of variance using the IBM SPSS Statistics 20 software (SPSS commercial software, USA).

Results

Screening for Optimal Trehalose Concentration

Under salt stress, the tomato leaves exhibited a withering state, and the plant height, fresh mass, and the relative water content (RWC) of the leaves were significantly reduced as shown in Figs. 2 and 3, respectively. However, exogenous trehalose improved the growth of tomato and increased the plant height, fresh mass, and the relative water content of tomato leaves, especially under treatment with 2 mmol/L trehalose, which was consistent with the results in Ref. [27].

Fig. 2
figure 2

Tomato leaves grown under normal and salt-treated conditions

Full size image
Fig. 3
figure 3

Effect of trehalose concentration on tomato growth under salt stress. A Plant height, B fresh mass, and C relative water content

Full size image

Salt stress affects photosynthesis in plants. Salt stress can increase the number of free radicals in chloroplasts, thereby destroy the chlorophyll, and affect the photosynthesis of plants. After exogenous trehalose treatment, the reduction in chlorophyll content was improved and plant salt stress was alleviated [28]. The chlorophyll content in leaves decreased significantly after salt stress treatment (Fig. 4). However, after treatment with different concentrations of exogenous trehalose, the chlorophyll content increased initially and then decreased. The highest chlorophyll content was found to be with 2 mmol/L trehalose treatment.

Fig. 4
figure 4

Effect of trehalose concentration on the chlorophyll content of tomato leaves under salt stress. A Chlorophyll a, B chlorophyll b, and C total chlorophyll

Full size image

The MDA content increased significantly after salt treatment as shown in Fig. 5A, indicating that salt stress destroyed the cell membrane of plants severely, and a large number of free radicals and reactive oxygen species were produced that resulted in membrane damage. After treatment with different concentrations of trehalose, the MDA content decreased initially and then increased and was significantly reduced with 2 mmol/L trehalose treatment.

Fig. 5
figure 5

Effects of different concentrations of trehalose on physiological indexes of tomato leaves under salt stress. A MDA, B proline, C CAT, D SOD, and E POD

Full size image

The accumulation of proline increases the osmotic potential of cells, regulates osmosis, and alleviates salt stress. Under salt treatment, proline showed a certain accumulation; after the application of trehalose, the proline content increased significantly, with the highest content being observed with 2 mmol/L trehalose treatment (Fig. 5B).

High salt content causes an imbalance of reactive oxygen species in plants, and plants remove the excess reactive oxygen species primarily through antioxidant enzymes such as SOD, POD, CAT, and other antioxidant enzymes and non-antioxidant enzymes such as ascorbic acid, glutathione (GSH), β-carotene, and mannitol [21]. It has been reported that the activities of SOD, CAT, and POD show a positive correlation with plant salt resistance [29]. In our study, the activities of these three antioxidant enzymes (POD, SOD, and CAT) increased after salt treatment, which indicated that salt stress could enhance the activities of these enzymes. After treatment with different concentrations of trehalose, the activities of the three antioxidant enzymes increased initially and then decreased, with the highest activity of CAT and SOD being detected with 2 mmol/L trehalose treatment (Fig. 5C, D). And the activity of POD reached the second highest, close to the highest, with 2 mmol/L trehalose treatment (Fig. 5E).

The TOPSIS mathematical model was used to analyze and screen the optimum exogenous trehalose concentration for alleviating salt stress in tomato seedlings. Results showed that the optimum concentration of trehalose was 2 mmol/L (Table 4). Therefore, this concentration was selected for subsequent experiments.

Table 4 Appropriate treatment concentration evaluated by the TOPSIS method
Full size table

Effect of Exogenous Trehalose on Sugar Content of Tomato Seedlings Under Salt Stress

The accumulation of starch can lead to a negative feedback mechanism of photosynthesis, reduce the intensity of photosynthesis, and slow down plant growth. Under salt stress, starch accumulated in tomato leaves and reached the maximum content on the 7th day, whereas the addition of trehalose reduced starch accumulation, as shown in Fig. 6. The changes in the starch content of roots were consistent with those of leaves, but the starch content in roots was lower than that in leaves, which may be due to the fact that salt stress destroyed the transport mechanism of starch from leaves to roots and affected starch transport, thus primarily accumulating in leaves and causing a negative feedback mechanism of photosynthesis.

Fig. 6
figure 6

Effect of exogenous trehalose on the starch content of tomato seedlings under salt stress. A Roots and B leaves

Full size image

Plants regulate the cell osmotic potential by absorbing external inorganic salt ions and synthesizing small organic molecules such as proline and soluble sugar (sucrose, trehalose, etc.) to resist salt stress [30, 31]. Soluble sugar can be used as an osmotic substance to alleviate the ionic stress of plants and as a signal molecule to stimulate the response to salt stress. Our study showed that under salt stress, the soluble sugar contents increased in tomato roots and leaves, and exogenous trehalose significantly increased the soluble sugar contents, especially in the later stage (Fig. 7). It may be due to the fact that trehalose reduced the accumulation of starch and promoted the conversion of starch into soluble sugar.

Fig. 7
figure 7

Effect of exogenous trehalose on the soluble sugar content of tomato seedlings under salt stress. A Roots and B leaves

Full size image

As shown in Fig. 8, exogenous trehalose treatment under salt stress increased sucrose content, which is the primary product of photosynthesis. Exogenous trehalose contributed to the accumulation of sucrose, indicating that it may induce an increase in photosynthesis rate, which was consistent with the abovementioned results showing that exogenous trehalose increased the content of photosynthetic pigments. The change in fructose levels was inconsistent compared with sucrose levels. Under salt stress, the fructose content increased, whereas exogenous trehalose treatment significantly decreased the fructose content. It was probably because trehalose reduced the hydrolysis rate of sucrose, and the rate of sucrose synthesis was greater than the rate of degradation. With the progression of salt treatment, the sorbitol content increased gradually. Exogenous trehalose further induced an increase in sorbitol content in the later stage.

Fig. 8
figure 8

Effects of exogenous trehalose on the contents of sucrose, fructose, and sorbitol in tomato seedlings under salt stress. A Sucrose in roots, B sucrose in leaves, C fructose in roots, D fructose in leaves, E sorbitol in roots, and F sorbitol in leaves

Full size image

In general, the content of trehalose is extremely low in plants, and it is difficult to detect its presence directly. As shown in Fig. 9, the trehalose content under normal conditions was very low, and salt stress stimulated trehalose production, especially in leaves. The increase in trehalose content confirmed that it enhanced the salt tolerance of tomato to some extent.

Fig. 9
figure 9

Effect of exogenous trehalose on the trehalose content of tomato seedlings under salt stress. A Roots and B leaves

Full size image

Effect of Exogenous Trehalose on Key Genes Involved in Sugar Metabolism in Tomato Seedlings Under Salt Stress

Starch is the primary product of plant photosynthesis and is generally degraded to resist stress. The starch synthase (SS) gene and the adenosine diphosphate glucose pyrophosphate (AGPase) gene play an important role in the starch synthesis pathway, while alpha amylase (AMY) and beta amylase (BAM) are the two key enzymes involved in plant starch degradation.

In our study, SS expression in tomato roots decreased initially and then increased under salt stress (Fig. 10). On the 1st day, exogenous trehalose significantly inhibited the expression of SS, which was consistent with the decrease in starch content. However, with the progression of salt treatment, SS expression gradually became stable. The change in the expression of SS in tomato leaves was approximately similar to that in roots. Under salt stress, AGPase expression in tomato roots was inhibited, especially in the case of exogenous trehalose treatment, which decreased its expression to 22.3% on the 5th day. However, in leaves, AGPase expression was increased significantly on the 1st day and then decreased with the progression of salt treatment.

Fig. 10
figure 10

Effect of exogenous trehalose on the expression of SS and AGPase in tomato seedlings under salt stress. ASS in roots, BSS in leaves, CAGPase in roots, and DAGPase in leaves

Full size image

As shown in Fig. 11, beta amylase played an important role in response to salt stress. The expression of the gene BAM in tomato roots was significantly upregulated compared with the control group, especially under treatment with exogenous trehalose (except for the 5th day), which promoted the degradation of starch and reduced the feedback inhibition of photosynthesis. In leaves, BAM expression increased initially and then decreased, which was basically consistent with the previous changes in starch content. Salt + trehalose treatment significantly upregulated the expression of the α-amylase gene AMY, with the highest expression being observed on the 3rd day in roots, which was approximately eight times of that in the control group and two times of that in the group of salt treatment alone, indicating that AMY was sensitive to trehalose. In leaves, the expression of AMY reached the peak on the 1st day and then decreased. The differential expression of AMY in different tissues indicated that its contribution to starch degradation was inconsistent and tissue specific.

Fig. 11
figure 11

Effect of exogenous trehalose on the expression of BAM and AMY in tomato seedlings under salt stress. ABAM in roots, BBAM in leaves, CAMY in roots, and DAMY in leaves

Full size image

Sucrose constitutes the highest proportion of sugars in plants and is directly related to plant stress resistance. The sucrose phosphate synthase (SPS) gene, the sucrose synthase (SUS3) gene, and the acid invertase (Wiv-1) gene are important in the sucrose metabolism pathway.

Figure 12 shows that SPS expression increased with the addition of exogenous trehalose under salt stress. Especially in roots, SPS expression was 14 times higher than that in the control group after 1 day of treatment, indicating that the effect of exogenous trehalose on roots was more obvious, which was consistent with the theory that roots are the most sensitive organs under salt stress. Sucrose synthase not only hydrolyzes sucrose but also synthesizes sucrose, regulating a reversible reaction. In roots, the expression of SUS3 was not significantly different from that in the control group and tended to be stable. In leaves, SUS3 expression decreased initially and then increased compared with that in the control group.

Fig. 12
figure 12

Effect of exogenous trehalose on the expressions of SPS, SUS3, and Wiv-1 in tomato seedlings under salt stress. ASPS in roots, BSPS in leaves, CSUS3 in roots, DSUS3 in leaves, EWiv-1 in roots, and FWiv-1 in leaves

Full size image

The enzyme acid invertase facilitates the hydrolysis of sucrose into glucose and fructose, which can significantly increase the cell osmotic potential. Salt treatment induced the expression of Wiv-1 in roots and leaves. Especially on the 3rd day, Wiv-1 expression reached the peak with salt + trehalose treatment, which was upregulated by 7.26 times and 10.68 times, respectively, indicating that Wiv-1 may play an essential role in salt stress, while exogenous trehalose treatment significantly stimulated its expression.

The trehalose phosphate synthase (TPS) gene, the trehalose phosphatase (TPP) gene, and the trehalose (Tre) gene play an important role in the synthesis and metabolism of trehalose. As shown in Fig. 13, salt stress induced the expression of the TPS gene in roots and leaves, and the upregulation was higher with salt + trehalose treatment, which was consistent with the trend of trehalose content in Fig. 9. The expression of TPP was significantly upregulated under salt stress, reaching peaks on the 3rd and 1st day in roots and leaves, respectively. Exogenous trehalose treatment inhibited the expression of this gene, which may be caused due to the negative feedback regulation. The expression of Tre in roots and leaves increased initially and then decreased and was enhanced by the addition of trehalose and yielded a peak after 1 day.

Fig. 13
figure 13

Effect of exogenous trehalose on the expression of TPS, TPP, and Tre in tomato seedlings under salt stress. ATPS in roots, BTPS in leaves, CTPP in roots, DTPP in leaves, ETre in roots, and FTre in leaves

Full size image

Effects of Exogenous Trehalose on Sugar Signal Transduction Genes and Sugar Transporter Genes of Tomato Seedlings Under Salt Stress

Sucrose is converted into fructose under the action of sucrose synthase and hydrolase, and fructose enters the glycolysis cycle under the action of fructose phosphokinase and phosphokinase, participating in plant stress response. Under salt stress, the expression of fructokinase (FRK2) gene decreased initially and then increased in roots, whereas salt + trehalose treatment significantly inhibited its expression, which began to be upregulated on the 7th day (Fig. 14). In leaves, the expression of FRK2 increased gradually under salt + trehalose treatment, reaching its peak on the 5th day.

Fig. 14
figure 14

Effect of exogenous trehalose on the expression of FRK2 and PFK in tomato seedlings under salt stress. AFRK2 in roots, BFRK2 in leaves, CPFK in roots, and DPFK in leaves

Full size image

As shown in Fig. 14C, D, salt stress significantly induced the expression of the phosphate fructokinase (PFK) gene, indicating that PFK may play an important role in response to salt stress. Moreover, salt + trehalose treatment further increased PFK expression in roots, which was upregulated by 9.3 times compared with that in the control group on the 1st day. PFK expression in tomato leaves was lower than that in roots but significantly upregulated.

Sucrose and hexose signal receptors are the major sugar signal receptors in plants. We selected two hexokinase genes, HXK2 and HXK3, for this study (Fig. 15). Both genes were upregulated to varying degrees under salt stress, and exogenous trehalose was more conducive to the expression of hexokinase gene. Under salt + trehalose treatment, the expression of HXK2 increased initially and then decreased, reaching its peak on the 3rd day. HXK3 expression was upregulated with both salt treatment and salt + trehalose treatment.

Fig. 15
figure 15

Effect of exogenous trehalose on the expression of HXK2 and HXK3 in tomato seedlings under salt stress. AHXK2 in roots, BHXK2 in leaves, CHXK3 in roots, and DHXK3 in leaves

Full size image

The transport and distribution of sugar directly affect the normal growth and development of plants and also play an important role in plant stress. We analyzed the changes in the expression of the monosaccharide transporter (MST3) gene and the sucrose transporter genes (SUT1 and SUT4) under salt stress. As shown in Fig. 16, the expression of MST3 in roots was upregulated with salt treatment and significantly upregulated with salt + trehalose treatment. MST3 expression in leaves was also induced by salt stress and reached the peak on the 1st day after salt + trehalose treatment. These results showed that both SUT1 and SUT4 were upregulated with salt + trehalose treatment, suggesting that these genes may be involved in the response to salt stress.

Fig. 16
figure 16

Effect of exogenous trehalose on the expression of MST3, SUT1, and SUT4 in tomato seedlings under salt stress. AMST3 in roots, BMST3 in leaves, CSUT1 in roots, DSUT1 in leaves, ESUT4 in roots, and FSUT4 in leaves

Full size image

Effects of Exogenous Trehalose on ABA Synthesis and Metabolism of Tomato Seedlings Under Salt Stress

Under stress conditions, there is an increase in the ABA content in plants to resist external stress. As shown in Fig. 17, the ABA content increased significantly under salt stress, indicating that ABA was a stress-related hormone. After salt treatment, exogenous trehalose significantly increased the ABA content in tomato roots and leaves, reaching their peaks on the 5th day.

Fig. 17
figure 17

Effect of exogenous trehalose on the ABA content in tomato seedlings under salt stress. A Roots and B leaves

Full size image

NCED1 and NCED2 are key genes in the ABA synthesis pathway. As shown in Fig. 18, NCED1 and NCED2 in tomato roots and leaves were upregulated under salt treatment, especially when exogenous trehalose was added, indicating that exogenous trehalose had a significant positive effect on ABA synthesis.

Fig. 18
figure 18

Effect of exogenous trehalose on the expression of NCED1 and NCED2 in tomato seedlings under salt stress. ANCED1 in roots, BNCED1 in leaves, CNCED2 in roots, and DNCED2 in leaves

Full size image

CYP707A1 and CYP707A2 are important genes in the ABA metabolic pathway. Figure 19 shows that the expression of CYP707A1 was upregulated under salt stress, especially in roots. When treated for 3 days, CYP707A1 expression was 1.62 times that of the control group. However, the addition of exogenous trehalose reduced the upregulated expression of CYP707A1 under salt stress. CYP707A2 expression was decreased, and exogenous trehalose further reduced its expression.

Fig. 19
figure 19

Effect of exogenous trehalose on the expression of CYP707A1 and CYP707A2 in tomato seedlings under salt stress. ACYP707A1 in roots, BCYP707A1 in leaves, CCYP707A2 in roots, and DCYP707A2 in leaves

Full size image

Discussion

Salt treatment inhibited the growth of tomato seedlings. After the application of trehalose, the plant damage was alleviated, whereas high concentrations of trehalose inhibited the growth of tomato seedlings. Although trehalose can provide a source of carbon for tomato, due to the inability of tomato to utilize carbon source under salt stress, ionic stress is aggravated, which reduces photosynthesis and affects tomato growth.

ROS accumulation increases significantly under salt stress, which may lead to cell membrane damage, oxidative stress, or cell death [32, 33]. In this study, we demonstrated that trehalose exerted the effects of enhancing the activities of antioxidant enzymes, increasing the proline content, and decreasing the MDA content, therefore alleviating the damage caused due to salt stress.

Under salt stress, starch accumulated in tomato roots and leaves, leading to a negative feedback mechanism of photosynthesis, which reduced the photosynthetic rate, decreased the chlorophyll content, and hindered starch transport. Exogenous trehalose treatment inhibited the expression of the starch synthesis genes SS and AGPase and promoted the expression of the starch metabolism genes BAM and AMY, thereby reducing the starch content, alleviating the negative feedback mechanism, and facilitating the accumulation of photoassimilation products.

Soluble sugars are primarily used as osmotic agents to prevent plant cells from dehydration and death and to maintain the internal stability of the cells under salt stress [34, 35]. When treated with salt, the content of various soluble sugars in different tissues of tomato seedlings increased significantly, while exogenous trehalose further increased their contents, especially that of sucrose. As an important osmotic regulator, sucrose can significantly increase the osmotic potential of plant cells and alleviate ionic stress. In addition, sucrose can eliminate ROS in plants. Exogenous trehalose administration significantly induced the expression of the SPS under salt stress and increased the sucrose content in tomato. The expression of the sucrose synthase gene SUS3 increased initially and then decreased, whereas that of the acid invertase gene Wiv-1 increased, which maintained a relative stability of sucrose content under salt stress.

The expression of the TPS in tomato was upregulated under salt stress, which was consistent with the findings of Li et al. [36] who showed that overexpression of OsTPS1 enhanced the salt tolerance of rice and exogenous trehalose treatment further increased TPS expression. In our experiment, exogenous trehalose administration increased the content of endogenous trehalose in tomato, resulted in a negative feedback regulation, inhibited the expression of the TPP, enhanced the expression of trehalose gene, and promoted the conversion of excessive trehalose into glucose to maintain the stability of its content. TPS, TPP, and Tre exhibited differential expressions under salt stress, suggesting that the trehalose metabolic pathway could directly affect the salt tolerance of plants.

Fructose phosphokinase and fructokinase are two important enzymes in the plant glycolytic pathway, which can maintain the normal physiological function of plants and alleviate the physiological damage occurring in plants under stress [37]. The results of this study showed that the addition of exogenous trehalose could significantly upregulate the expressions of FRK and PFK under salt stress, suggesting that it could help plants accumulate sugars and maintain strong respiration. Sugar not only is the primary energy source for plant growth, but also can be used as a signal molecule to respond to plant stress. HXK is a sugar receptor that is affected by the external environment and forms a network signal system with plant hormones such as ethylene, ABA, and gibberellin. Exogenous trehalose treatment induced the upregulation of HXK2 and HXK3 to different degrees under salt stress. Sami et al. [38] reported that sugar and ABA interacted in the metabolic pathways of plants to influence drought and salt resistance.

Salt stress affects the activities of sugar transporters on plant membranes, which results in the abnormal transport of sugar and the inability of plants to obtain energy [39]. The sugar transporter genes exhibited different expression trends during osmotic stress, suggesting that the source–sink relationship of carbon may be related to the regulation of sugar transporter gene expression [40]. In addition, AtSUC4 could regulate sucrose distribution and metabolism in response to drought stress [41]. The results of this study demonstrated that the expressions of the monosaccharide transporter (MST3) gene and the sucrose transporter genes SUT1 and SUT4 were significantly upregulated with salt + trehalose treatment. Trehalose may change the source–sink transport pathway of sugars, thereby changing the accumulation and distribution of sugars to respond to salt stress.

The entire process of plant growth and development is affected by the interaction between sugar and ABA. In this experiment, the ABA content of tomato seedlings accumulated significantly under salt stress, which was consistent with the results of Kong et al. [42] and Shu et al. [43], indicating that ABA directly responded to plant salt stress. Salt + trehalose treatment resulted in the upregulation of the ABA synthesis-related genes and the downregulation of metabolic genes, which indicated that the tomato plant increased its ABA content by accelerating ABA synthesis and reducing ABA decomposition. Saeedipour [44] found that differences in salt stress tolerance of rice plant were related to its ABA synthesis ability under stress conditions. Tomatoes overexpressing NCED accumulated more ABA and showed improved drought resistance. ABA primarily improves plant stress resistance by closing the stomata of plant leaves, reducing transpiration, and maintaining water in plants [45]. Therefore, trehalose acts as a signaling molecule by affecting the ABA anabolic pathway, leading to ABA accumulation and further improving the salt tolerance of tomato.

In summary, trehalose, a soluble, non-reducing disaccharide, is present in a low amount in plants; however, it regulates sugar accumulation and distribution by affecting the activities of sugar transporters and simultaneously regulates sugar metabolism and ABA metabolism, thereby significantly improving plant stress resistance (Figs. 20, 21). This study provides a greater understanding of the salt-tolerant mechanism of plants and has important value for improving crop yield, salt tolerance of plants, and utilization of saline–alkali land in agricultural research.

Fig. 20
figure 20

Changes in glucose metabolism in tomato seedlings under salt stress by exogenous trehalose

Full size image
Fig. 21
figure 21

Changes in ABA metabolism in tomato seedlings under salt stress by exogenous trehalose

Full size image

Future research will continue to explore the effect of trehalose on tomato salt stress using transgenic technology to alter the expression of genes related to trehalose metabolism, which could help in further studying the salt tolerance mechanism of trehalose.