Glutathione Transferases Are Involved in the Genotype-Specific Salt-Stress Response of Tomato Plants

Abstract

Glutathione transferases (GSTs) are one of the most versatile multigenic enzyme superfamilies. In our experiments, the involvement of the genotype-specific induction of GST genes and glutathione- or redox-related genes in pathways regulating salt-stress tolerance was examined in tomato cultivars (Solanum lycopersicum Moneymaker, Mobil, and Elán F1). The growth of the Mobil plants was adversely affected during salt stress (100 mM of NaCl), which might be the result of lowered glutathione and ascorbate levels, a more positive glutathione redox potential (E_GSH), and reduced glutathione reductase (GR) and GST activities. In contrast, the Moneymaker and Elán F1 cultivars were able to restore their growth and exhibited higher GR and inducible GST activities, as well as elevated, non-enzymatic antioxidant levels, indicating their enhanced salt tolerance. Furthermore, the expression patterns of GR, selected GST, and transcription factor genes differed significantly among the three cultivars, highlighting the distinct regulatory mechanisms of the tomato genotypes during salt stress. The correlations between E_GSH and gene expression data revealed several robust, cultivar-specific associations, underscoring the complexity of the stress response mechanism in tomatoes. Our results support the cultivar-specific roles of distinct GST genes during the salt-stress response, which, along with WRKY3, WRKY72, DREB1, and DREB2, are important players in shaping the redox status and the development of a more efficient stress tolerance in tomatoes.

Keywords: Solanum lycopersicum L.; glutathione transferases; redox homeostasis; salt stress; transcription regulation.

Figure 1

Growth of tomato plants under salt stress. Length and fresh weight of shoots (A,C) and roots (B,D) of five-week-old tomato plants measured under control conditions and after one week of 100 mM NaCl treatment. Mm—cv. Moneymaker, Mo—cv. Mobil, El—cv. Elán F1. Means ± SD. Significant differences according to the Student’s t-test (* p ≤ 0.05, ** p ≤ 0.01) are indicated.

Figure 2

Hydrogen peroxide (H₂O₂) levels (A,B) and malondialdehyde content (C,D) of five-week-old tomato plants under control conditions and after one week of 100 mM NaCl treatment. Mm—cv. Moneymaker, Mo—cv. Mobil, El—cv. Elán F1. Means ± SD. Significant differences according to the Student’s t-test (*** p ≤ 0.001) are indicated.

Figure 3

Ascorbic acid (reduced—AsA, oxidized—DHA; (A,B)) and glutathione (reduced—GSH, oxidized—GSSG; (C,D)) contents and their ratios in five-week-old tomato cultivars under control conditions and after one week of 100 mM NaCl treatment. Note the different vertical scales on the leaves (A,C) and roots (B,D). Mm—cv. Moneymaker, Mo—cv. Mobil, El—cv. Elán F1. Means ± SD. Significant differences according to the Student’s t-test (* p ≤ 0.05, ** p ≤ 0.01, *** p ≤ 0.001) are indicated.

Figure 4

Specific dehydroascorbate reductase (DHAR), glutathione reductase (GR), transferase (GST), and peroxidase (GPOX) activities of tomato cultivars in five-week-old tomato leaves (A,C,E,G) and roots (B,D,F,H), respectively, under control conditions and after one week of 100 mM NaCl treatment. Mm—Moneymaker, Mo—Mobil, El—Elán F1. Means ± SD. Significant differences according to the Student’s t-test (* p ≤ 0.05, ** p ≤ 0.01, *** p ≤ 0.001) are indicated.

Figure 5

Heat map of the expression levels of 2 Solanum lycopersicum glutathione reductases (GR1 and GR2) and 27 selected genes belonging to 4 GST classes (DHAR, GSTF, GSTU, and GSTT) determined in salt-treated, five-week-old Moneymaker (Mm), Mobil (Mo), and Elán F1 (El) tomato cultivars. The relative transcript amounts of the genes in the leaves (A) and roots (B) were determined by HT-qPCR after one week of 100 mM NaCl treatment. The expression of the genes was normalized first by the average of actin2 and elongation factor 1α genes, and second by the average transcript amount of each gene in untreated cultivars. The log₂ transformation of 2^−ΔΔCt data are presented. The green color represents repression, while the red color represents activation, as indicated on the color scale bar. The presented data are from two biological replicates.

Figure 6

Correlation analysis between the glutathione redox potential (E_GSH) and the expression of selected genes (ΔCt values), measured under different conditions in the leaves (A) and roots (B) of Moneymaker (Mm), Mobil (Mo), and Elán F1 (El) tomato plants. Positive correlations are highlighted in red and negative correlations are highlighted in blue.

Figure 7

Predicted cis regulatory elements found in the upstream regulatory region of GSTF2, GSTU15, GSTU32, and GSTU47 using the NewPLACE database (https://www.dna.affrc.go.jp/PLACE/?action=newplace, accessed on 25 February 2023).

Figure 8

Heat map of the expression levels of eight Solanum lycopersicum transcription factor genes. The relative transcript levels of the selected genes were determined in five-week-old tomato (Moneymaker—Mm, Mobil—Mo, and Elán F1—El) leaves (A) and roots (B) by qPCR after one week of 100 mM NaCl treatment. For this, the expression values were normalized, firstly by the average expression of the actin2 gene, and secondly by the average transcript levels of each gene from the untreated cultivars. The log₂ transformation of the 2^−ΔΔCt data are presented as a heat map. The green color represents repression, while the red color represents activation, as indicated on the color scale bar. The presented data have been obtained from two biological replicates. n.d.—not detected.

Figure 9

Correlation analysis on the basis of the glutathione redox potential (E_GSH) and expression of selected genes (ΔCt values), measured under different conditions in the leaves (A) and roots (B) of Moneymaker (Mm), Mobil (Mo), and Elán F1 (El) tomato plants. Red color highlights the positive correlations and the blue color highlights the negative correlations. n.d.—not detected.

Figure 10

Schematic model summarizing the potential regulators of GST gene expression after one week 100 mM NaCl treatments in tomato plants, based on our results and data found in the literature [90,91]. The salt treatment supposedly disturbed the redox homeostasis, which led to a temporary oxidized redox state within the cells. These changes in the redox potential may have had a direct effect on the expression of GSTs, but also led to the induction of the TF genes, such as WRKY3, WRKY72, DREB1, and DREB2. Presumably, the expression of these genes was also influenced by hormonal changes or, for example, 5′ cis regulatory elements could also induce the expression of GSTU15, GSTU32, and GSTU47 genes in all investigated cultivars. Abbreviations: Moneymaker—Mm; Mobil—Mo; Elán F1—El; control—C; salt—one week of 100 mM NaCl treatment.