Deciphering vital transcription factors in cytokinin- and ABA-mediated postharvest leaf senescence of Brassica rapa

Leaf senescence after harvest limits the economic value of leafy cruciferous vegetables such as Chinese flowering cabbage (Brassica rapa ssp. parachinensis). Understanding the intricate gene regulatory networks that govern postharvest leaf senescence offers potential strategies to extend the shelf life of these vegetables. This study elucidated the regulatory networks modulating leaf senescence by utilizing time-series gene expression analysis on postharvest leaves of Chinese flowering cabbage treated with cytokinin analog 6-benzylaminopurine and abscisic acid (ABA). ABA treatment accelerated leaf senescence, including the dismantling of chloroplasts and mitochondria, whereas 6-benzylaminopurine treatment decelerated these processes. Subsequent RNA sequencing and integrated analyses led to the construction of transcriptional regulatory networks comprising 49 transcription factors potentially regulating senescence-related pathways, including reactive oxygen species (ROS) metabolism and chlorophyll degradation. Validation experiments on ROS metabolism confirmed that increased ROS accumulation paralleled the progression of leaf senescence, whereas ABA and 6-benzylaminopurine treatment resulted in opposing effects on ROS scavenging. Furthermore, exogenous ROS treatment promoted leaf senescence and the disassembly of chloroplasts and mitochondria, while ROS inhibitors delayed these processes. Further validation assays affirmed the expression patterns, transcription factor-binding capacities, and activation potentials of eight critical transcription factors and their possible target genes associated with ROS scavenging. Moreover, the role of two transcription factors (BrAGL42 and BrCRF11-2) in regulating postharvest leaf senescence and ROS scavenging ability was verified through transformation assays. Collectively, our findings shed light on the overarching transcription factor-mediated regulatory pathways in postharvest leaf senescence and indicate how cytokinin and ABA modulate this process antagonistically.