Exogenous Putrescine Increases Heat Tolerance in Tomato Seedlings by Regulating Chlorophyll Metabolism and Enhancing Antioxidant Defense Efficiency

Abstract

Crops around the world are facing a diversity of environmental problems, of which high temperatures are proving to be the most serious threat to crops. Polyamine putrescine (Put) acts as a master growth regulator that contributes to optimal plant growth and development and increased stress tolerance. Here, the current study aimed to elucidate how Put functions in regulating chlorophyll (Chl) metabolism, oxidative stress, and antioxidant defense, as well as to characterize the expression of genes related to heat stress in tomato seedlings under such stress. The results revealed that Put treatment significantly attenuates heat-induced damage by promoting biomass production, increasing photosynthetic efficiency, and inhibiting excessive production of oxidative stress markers. Heat stress markedly decreased the Chl content in the tomato leaf and accelerated the leaf yellowing process. However, Put-treated tomato seedlings showed a higher Chl content, which could be associated with the functions of Put in elevating PBGD activity (Chl biosynthesis enzyme) and suppressing the activity of the Chl catabolic enzyme (Chlase and MDCase). Under high-temperature stress, the expression levels of the gene encoding factors involved in Chl biosynthesis and Chl catabolism were significantly down- and upregulated, respectively, and this trend was reversed in Put-treated heat-stressed seedlings. In addition, exogenous application of Put boosted the activity of antioxidant enzymes, along with the levels of expression of their encoding genes, only in plants that were heat stressed. Furthermore, the expression levels of heat-shock-related genes (HSP90, HSP70, and HsfA1) were elevated in Put-treated, high-temperature-stressed tomato seedlings. Taken together, our results indicate that Put treatment significantly increases the heat tolerance of tomato seedlings, by elevating Chl concentrations and suppressing Chl catabolic enzyme activity, modulating endogenous free PA content, increasing antioxidant defense efficiency, and upregulating the expression of heat-shock-related genes.

Keywords: chlorophyll degradation; oxidative stress; photosynthesis; polyamines; thermotolerance; tomato.

Figure 1

Interactive effects of Putrescine (Put) and high temperature on Chlorophyll (Chl) and carotenoid (Car) content in tomato seedlings. (A) Chl a content, (B) Chl b content, (C) Total Chl content, and (D) Car content. The data denote the mean value ± standard error (n = 3). Different alphabetic letters represent the significant differences among the treatments at p < 0.05, according to Tukey’s test.

Figure 2

Interactive effects of Putrescine and high temperature on (A) δ-Aminolevulinic acid (ALA) content, (B) Porphobilinogen (PBG) content, (C) Chlorophyllase (Chlase) activity, (D) Porphobilinogen deaminase (PBGD) activity, and (E) Mg-dechelatase (MDCase) activity in tomato seedlings. The data denote the mean value ± standard error (n = 3). Different alphabetic letters represent the significant differences among the treatments at p < 0.05, according to Tukey’s test.

Figure 3

Interactive effects of Putrescine and high temperature on Chlorophyll related genes expression in tomato seedlings. (A) Porphobilinogen deaminase (PBGD), (B) Mg-chelatase (Mg-CHT), (C) Chl synthase (CHL G), (D) Chlorophyllide a oxygenase (CAO) activity, (E) protochlorophyllide oxidoreductase (POR), (F) Chlorophyllase (CHLASE), (G) nonyellow coloring (NYC1), (H) pheophorbide a oxidase (PAO), and (I) stay-green (SGR). The data denote the mean value ± standard error (n = 3). Different alphabetic letters represent the significant differences among the treatments at p < 0.05, according to Tukey’s test.

Figure 4

Interactive effects of Putrescine and high temperature on gas exchange parameter content in tomato seedlings. (A) Net photosynthetic rate (Pn) content, (B) Stomatal conductance (Gs) content, (C) Intercellular carbon dioxide (CO₂) concentration (Ci) content, and (D) Transpiration rate (Tr) content. The data denote the mean value ± standard error (n = 3). Different alphabetic letters represent the significant differences among the treatments at p < 0.05, according to Tukey’s test.

Figure 5

Interactive effects of Putrescine and high temperature on photosynthetic attributes in tomato seedlings. (A) Maximum quantum yield of PSII (Fv/Fm), (B) Effective quantum efficiency of PS II (Y(II)), (C) Photochemical quenching coefficient (qP), and (D) Non-photochemical quenching (NPQ). The data denote the mean value ± standard error (n = 3). Different alphabetic letters represent the significant differences among the treatments at p < 0.05, according to Tukey’s test.

Figure 6

Interactive effects of Putrescine and high temperature on (A) Proline content, (B) Soluble sugar content, (C) Electrolyte leakage, and (D) MDA content in tomato seedlings. The data denote the mean value ± standard error (n = 3). Different alphabetic letters represent the significant differences among the treatments at p < 0.05, according to Tukey’s test.

Figure 7

Interactive effects of Putrescine and high temperature on (A) Accumulation of hydrogen peroxide, (B) Hydrogen peroxide content, (C) Accumulation of superoxide anion, and (D) Superoxide anion production rate in tomato seedlings. The data denote the mean value ± standard error (n = 3). Different alphabetic letters represent significant differences among the treatments at p < 0.05, according to Tukey’s test.

Figure 8

Interactive effects of Putrescine and high temperature on antioxidant enzymes activity in tomato seedlings. (A) Superoxide dismutase (SOD) activity, (B) Catalase (CAT) activity, (C) Peroxidase (POD) activity, (D) Ascorbate peroxidase (APX) activity, (E) Lipoxygenase (LOX) activity, (F) Glutathione S-transferase (GST) activity, (G) Glutathione reductase (GR) activity, (H) Monodehydroascorbate reductase (MDHAR) activity, and (I) Dehydroascorbate reductase (DHAR) activity. The data denote the mean value ± standard error (n = 3). Different alphabetic letters represent significant differences among the treatments at p < 0.05, according to Tukey’s test.

Figure 9

Heat map showing the expression of different stress-related genes in tomato leaves exposed to high temperature in the presence or absence of putrescine treatment. The intensity of gene expression ranges from deep green (low) color to deep red color (high). CK: control; Put: 1 mM putrescine; HT: heat stress (38/28 °C); Put + HT: 1 mM putrescine and heat stress. Antioxidant-related genes (FeSOD, MnSOD, CAT, POD, APX, GR, MDHAR, DHAR, GST, LOX); and Heat shock-related genes (HSP90, HSP70, HSfA1).

Figure 10

Interactive effects of Putrescine and high temperature on free polyamine content in tomato seedlings. (A) Free putrescine (Put) content, (B) Free spermidine (Spd) content, and (C) Free spermine (Spm) content. The data denote the mean value ± standard error (n = 3). Different alphabetic letters represent the significant differences among the treatments at p < 0.05, according to Tukey’s test.