In this research, the lettuce high-temperature-sensitive variety Beisan San 3 was used as a test material. The effects of exogenous spermidine (Spd) on membrane lipid peroxidation, the antioxidant system, the ascorbic acid-glutathione (AsA-GSH) system and the glyoxalase (Glo) system in lettuce seedlings under high-temperature stress were studied by spraying either 1 mM spermidine or ionized water as a control. The results showed that, under high-temperature stress, the growth of lettuce seedlings was weak, and the dry weight (DW) and fresh weight (FW) were reduced by 68.9% and 82%, respectively, compared with those of the normal-temperature controls. In addition, the degree of membrane lipid peroxidation increased, and the reactive oxygen species (ROS) level increased, both of which led to a significant increase in malondialdehyde (MDA) content and lipoxygenase (LOX) activity. Under high-temperature stress, the activity of superoxide dismutase (SOD) decreased, the activities of peroxidase (POD) and catalase (CAT) increased first but then decreased, and the activity of ascorbic acid peroxidase (APX) decreased first but then increased. Glutathione reductase (GR) activity, ascorbic acid (AsA) and glutathione (GSH) content showed an upward trend under high-temperature stress. The activities of glyoxalase (GloI and GloII) in the lettuce seedling leaves increased significantly under high-temperature stress. In contrast, the application of exogenous Spd alleviated the oxidative damage to the lettuce seedlings, which showed a decrease in MDA content and LOX activity and an increase in SOD, POD, CAT, APX, GR, GloI, and GloII activities. In addition, the antioxidant AsA and GSH contents also increased to varying degrees. It can be seen from the results that high temperature stress leads to an increase in the level of ROS and cause peroxidation in lettuce seedlings, and exogenous Spd can enhance the ability of lettuce seedlings to withstand high temperature by enhancing the antioxidant system, glyoxalase system and AsA-GSH cycle system.

Bolting is a key process in the growth and development of lettuce (Lactuca sativa L.). A high temperature can induce early bolting, which decreases both the quality and production of lettuce. However, knowledge of underlying lettuce bolting is still lacking. To better understand the molecular basis of bolting, a comparative proteomics analysis was conducted on lettuce stems, during the bolting period induced by a high temperature (33 °C) and a control temperature (20 °C) using iTRAQ-based proteomics, phenotypic measures, and biological verifications using qRT-PCR and Western blot. The high temperature induced lettuce bolting, while the control temperature did not. Of the 5454 identified proteins, 619 proteins presented differential abundance induced by high-temperature relative to the control group, of which 345 had an increased abundance and 274 had a decreased abundance. Proteins with an abundance level change were mainly enriched in pathways associated with photosynthesis and tryptophan metabolism involved in auxin (IAA) biosynthesis. Moreover, among the proteins with differential abundance, proteins associated with photosynthesis and tryptophan metabolism were increased. These findings indicate that a high temperature enhances the function of photosynthesis and IAA biosynthesis to promote the process of bolting, which is in line with the physiology and transcription level of IAA metabolism. Our data provide a first comprehensive dataset for gaining novel understanding of the molecular basis underlying lettuce bolting induced by high temperature. It is potentially important for further functional analysis and genetic manipulation for molecular breeding to breed new cultivars of lettuce to restrain early bolting, which is vital for improving vegetable quality.

The mitogen-activated protein kinase (MAPK) pathway is a widely distributed signaling cascade in eukaryotes and is involved in regulating plant growth, development, and stress responses. High temperature, a frequently occurring environmental stressor, causes premature bolting in lettuce with quality decline and yield loss. However, whether MAPKs play roles in thermally induced bolting remains poorly understood. In this study, 17 LsMAPK family members were identified from the lettuce genome. The physical and chemical properties, subcellular localization, chromosome localization, phylogeny, gene structure, family evolution, cis-acting elements, and phosphorylation sites of the LsMAPK gene family were evaluated via in silico analysis. According to phylogenetic relationships, LsMAPKs can be divided into four groups, A, B, C, and D, which is supported by analyses of gene structure and conserved domains. The collinearity analysis showed that there were 5 collinearity pairs among LsMAPKs, 8 with AtMAPKs, and 13 with SlMAPKs. The predicted cis-acting elements and potential phosphorylation sites were closely associated with hormones, stress resistance, growth, and development. Expression analysis showed that most LsMAPKs respond to high temperatures, among which LsMAPK4 is significantly and continuously upregulated upon heat treatments. Under heat stress, the stem length of the LsMAPK4-knockdown lines was significantly shorter than that of the control plants, and the microscope observations demonstrated that the differentiation time of flower buds at the stem apex was delayed accordingly. Therefore, silencing of LsMAPK4 significantly inhibited the high- temperature-accelerated bolting in lettuce, indicating that LsMPAK4 might be a potential regulator of lettuce bolting. This study provides a theoretical basis for a better understanding of the molecular mechanisms underlying the MAPK genes in high-temperature-induced bolting.

High temperature is a huge threat to lettuce production in the world, and spermidine (Spd) has been shown to improve heat tolerance in lettuce, but the action mechanism of Spd and the role of polyamine metabolism are still unclear. The effects of Spd and D-arginine (D-arg) on hydroponic lettuce seedlings under high-temperature stress by foliar spraying of Spd and D-arg were investigated. The results showed that high-temperature stress significantly inhibited the growth of lettuce seedlings, with a 33% decrease in total fresh weight and total dry weight; photosynthesis of lettuce seedlings was inhibited by high-temperature stress, and the inhibition was greater in the D-arg treatment, while the Spd recovery treatment increased net photosynthetic rate (Pn), stomatal conductance (Gs), transpiration rate (Tr), stomatal limit value (Ls), and intercellular CO₂ concentration (Ci). High-temperature stress significantly reduced the maximum photochemical efficiency (Fv/Fm), photochemical quenching coefficient (qP), electron transport rate (ETR), and photochemical efficiency of PSII (ΦPSII), increased the non-photochemical burst coefficient (NPQ) and reduced the use of light energy, which was alleviated by exogenous Spd. The increase in polyamine content may be due to an increase in polyamine synthase activity and a decrease in polyamine oxidase activity, as evidenced by changes in the expression levels of genes related to polyamine synthesis and metabolism enzymes. This evidence suggested that D-arg suppressed endogenous polyamine levels in lettuce and reduced its tolerance, whereas exogenous Spd promoted the synthesis and accumulation of polyamines in lettuce and increased its photosynthetic and oxidative stress levels, which had an impact on the tolerance of lettuce seedlings.