Single-cell and Spatial Transcriptomics Define 20E-Driven Developmental Reprogramming in Silkworm Wing Disc

Insect wing development involves tissue patterning, cell fate transitions, and hormone signaling, yet its spatiotemporal logic remains unclear. The silkworm, with large wing discs and defined stages, provides an ideal model for high resolution analysis. Here, we construct a spatiotemporal single-cell atlas of the silkworm wing disc across 10 timepoints, identifying 12 major cell types and their developmental transitions. Wing morphogenesis (Wm) cells act as central progenitors, differentiating into epithelial and cuticle lineages under lineage-specific transcription factors. Time‑resolved snRNA‑seq reveals hierarchical transcriptional reprogramming, with Wm cells functioning as early signaling hubs. Functional modules and signaling pathways were activated in spatiotemporal controlled manner. 20‑hydroxyecdysone treatment rapidly accelerates fate transitions and gene expression, recapitulating natural development within hours. Integration of morphology, hormone levels, and gene expression supports a five-stage Gene Transition Model describing progressive fate resolution. This work reveals wing development in silkworm and provides insights into hormone-driven organogenesis and potential manipulation of insect development in agriculture.