Nocturnal red light modulates the ethylene production and the downstream transcriptional networks that control plant defence responses at dawn

• Application of nocturnal red light altered leaf thickness and internode length of tomato. • Ethylene insensitivity influenced the expression of photosynthesis, circadian, and flowering regulators. • Red light caused salicylic- and abscisic acid accumulation in an ethylene-dependent manner. • Red light increased ascorbate and GST activity, which depended on ethylene. • Red light reduced B. cinerea infection independently of ET action. The interaction between ethylene (ET) and light/dark cycles is complex and regulates plant growth, development and stress responses. While ET synthesis is influenced by circadian rhythms, light -especially red light- it can also affect ET-regulated processes, depending on the developmental stage and organ type. This study investigated the effects of nocturnal red light exposure on mature tomato plant leaves, using wild-type (WT) and ET-insensitive Never ripe ( Nr ) mutant. Red light application for one week at night did not significantly affect leaf biomass and area, but it altered structural traits such as leaf thickness and internode length, particularly in WT plants. Transcriptomic analyses indicated significant differences in gene expression between WT and Nr plants, particularly in genes related to photosynthesis, growth regulation and the circadian clock. Red light also modulated the expression of hormone-related genes and increased the levels of defence-related phytohormones (ET, salicylic acid, abscisic acid) as well as the activity of antioxidant enzymes (e.g. guaiacol peroxidase, glutathione S-transferase) in a manner dependent on ET in WT plants. Interestingly, although ET is involved in many of these processes, red light was also found to increase biotic stress resistance in Nr mutants, as evidenced by reduced Botrytis cinerea infection. This suggests that red light may activate defence pathways independently of ET. Our findings highlight the potential of red light as an environmentally friendly tool for improving crop disease resistance and emphasise the pivotal role of ET signalling, which can be regulated by specific light spectra, in plant’s defence reactions.