Diel regulation in spinach synchronizes nitrogen assimilation via transcriptional gating

assimilation, chlorophyll content, and photosynthetic efficiency, with maxima in the middle of the light phase. Transcriptome analysis revealed that the greatest gene reprogramming occurred at the dark-to-light transition, where the expression of the morning oscillator component LHY peaked and coincided with the accumulation of dawn-biased nitrate-assimilation transcripts (NIA and NiR) and transporters. Peaks enzyme activities lagged transcript peaks by nearly one phase, suggesting transcriptional priming followed by daytime nitrogen assimilation, aligned with photosynthetic energy and carbon skeleton synthesis. Amino acid profiles reflected this coordination, with glutamine elevated during the day and glutamate enriched at night. Nitrogen deficiency suppressed growth, caused accumulation of reactive oxygen species, and activated circadian-regulated DNA repair genes (e.g., MSH2 and RPA2A, indicating genotoxic stress. Together, these findings support a model in which a diel/clock program, amplified by nitrogen sufficiency, gates nitrogen transport and reduction at dawn to couple daytime assimilation with photosynthesis while adjusting oxidative and DNA repair responses. This mechanistic framework clarifies how temporal regulation and nutrient status interact to shape nitrogen use efficiency and stress resilience in leafy vegetables.