Saline–alkali stress is a significant abiotic stressor that affects the growth of tomato (Solanum lycopersicum L.). Our previous studies reported that the homologous genes SlWRKY80 and SlWRKY81 can both resist saline-alkali stress by mediating the JA pathway. In this study, the co-overexpressing (SOE) and co-silencing (SQ) lines of SlWRKY80 and SlWRKY81 to saline-alkali treatment, and the results showed that SOE line had significantly higher morphological indices, physiological indices, and endogenous JA and MeJA contents than the WT, while SQ line was opposite. In addition, the SOE line showed a significant increase in autophagy against saline-alkali stress, and both SlWRKY80 and SlWRKY81 were positively regulating the transcription of the autophagy gene SlATG13b as screened and verified by transcriptome screening, qRT-PCR, WB (Western Blot), Y1H (Yeast One-Hybrid), ChIP-qPCR, EMSA (Electrophoretic Mobility Shift Assay), and D-LUC (D-Luciferin Experiment), and also had a synergistic effect in activating SlATG13b. The growth and physiological indexes of VIGS-SlATG13b line were significantly lower than those of other groups, and pro-SlATG13b is significantly activated by MeJA and saline-alkali signals, so the SlATG13b was regulating the resistance of tomato to saline-alkali stress. In summary, the increase of endogenous JA and MeJA content in tomato after saline-alkali stress inhibited the expression and synthesis of SlJAZ1, activated the transcription and protein synthesis of SlWRKY80 and SlWRKY81, and SlWRKY80 and SlWRKY81 coordinately regulated the transcription of SlATG13b, which resulted in the production of more autophagosomes and enhanced the resistance of tomato to saline-alkali stress. This study further improved the molecular mechanism of SlWRKY80 and SlWRKY81 mediating the JA pathway against saline-alkali stress, and laid a solid theoretical foundation for tomato saline-alkali resistance breeding.

The excess amounts of heavy metals are toxic to various physiological processes in plants. Plant metallothioneins (MTs) are low molecular weight, cysteine-rich metal-binding proteins that play important roles in the detoxification of heavy metal ions. In this study, we characterized CsMT4, a plant type 4 MT gene identified in a complementary DNA (cDNA) library prepared from young cucumber (Cucumis sativus) fruit. CsMT4 encodes a 90 amino acid protein with a predicted molecular mass of 9.028 kD. CsMT4 contains 17 cysteine residues in three highly conserved cysteine-rich domains. In contrast to the structures of other MTs, the highly conserved amino acid pattern CxCxxxCxCxxCxC is present in the middle of CsMT4. Furthermore, CsMT4 was markedly induced in various tissues by various concentrations of Cd²⁺ and Zn²⁺ (> 1.0 mmol•L⁻¹). Similar to AtMT4b function, heterologous expression of CsMT4 in E. coli could also improved its tolerance to Cd²⁺ and led to increased uptake of Cd²⁺, and the rate of Cd²⁺ uptake was the highest in cells expressing a phytochelatin-like peptide. Our findings demonstrate that CsMT4 might be obviously induced by the metal stress in cucumber, and improves tolerance to Cd ions but not Zn ions when heterologously expressed in E. coli, and suggest that the composition and arrangement of N-terminal Cys-residues in MT4 are associated with their binding capacity and preference for different metal ions.