Transcriptional landscapes of de novo root regeneration from detached Arabidopsis leaves revealed by time-lapse and single-cell RNA sequencing analyses.

Detached Arabidopsis thaliana leaves can regenerate adventitious roots, providing a platform for studying de novo root regeneration (DNRR). However, the comprehensive transcriptional framework of DNRR remains elusive. Here, we provide a high-resolution landscape of transcriptome reprogramming from wound response to root organogenesis in DNRR and show key factors involved in DNRR. Time-lapse RNA sequencing (RNA-seq) of the entire leaf within 12 h of leaf detachment revealed rapid activation of jasmonate, ethylene, and reactive oxygen species (ROS) pathways in response to wounding. Genetic analyses confirmed that ethylene and ROS may serve as wound signals to promote DNRR. Next, time-lapse RNA-seq within 5 d of leaf detachment revealed the activation of genes involved in organogenesis, wound-induced regeneration, and resource allocation in the wounded region of detached leaves during adventitious rooting. Genetic studies showed that BLADE-ON-PETIOLE1/2, which control aboveground organs, PLETHORA3/5/7, which control root organogenesis, and ETHYLENE RESPONSE FACTOR115, which controls wound-induced regeneration, are involved in DNRR. Furthermore, single-cell RNA-seq data revealed gene expression patterns in the wounded region of detached leaves during adventitious rooting. Overall, our study not only provides transcriptome tools but also reveals key factors involved in DNRR from detached Arabidopsis leaves.