Single-cell and Spatial Transcriptomics Reveals the Spatiotemporal Trajectory of the Small Peptide TAP4 in Delaying Postharvest Fruit Senescence

Trypsin has been reported to possess specific superoxide anion-scavenging activity and to exert pronounced anti-senescence effects in H. undatus fruit; however, the underlying mechanisms responsible for its anti-senescence function remain unclear. This study identified TAP4 as the most bioactive peptide among the trypsin autolysis peptides (TAPs). Single-cell transcriptomic analysis revealed that TAP4 alters pericarp cell differentiation trajectories, particularly in endocarp cells, during senescence. It activated the transcription factor HuWRKY21-1, specifically inducing immune responses in inner pericarp cells and upregulating 44 immune-related genes, including HuIGP4 and HuLYK6. Downstream factors such as FULL and ERFs promoted biosynthesis of antioxidants like antheraxanthin and hyperin. Functional validation via VIGS, callose deposition, disease resistance assays, and RT-qPCR confirmed the roles of key transcription factors such as HuWRKY21-1, HuFULL, and HuERF30-1 in immune activation and resistance reconstruction. This work provides evidence for the potential of the endocarp as defensive tissue in plant fruit and opens new avenues for understanding the mechanisms underlying the establishment of plant resistance.