A multi-layer regulatory landscape of chilling requirement revealed by stage-resolved transcriptomics and hormone profiling in peach floral buds

Bud dormancy and chilling requirement (CR) are fundamental to the climate adaptation of temperate fruit trees, yet the regulatory mechanisms translating winter chill into genotype-specific dormancy release remain poorly resolved. Here, we generated a stage-resolved transcriptomic and hormone dataset from peach floral buds of four cultivars spanning contrasting CR phenotypes, sampled across endodormancy induction (S1), maintenance (S2), and release (S3), with matched ABA/GA measurements. By applying trajectory-centric analysis that standardizes temporal profiles within each cultivar, we partitioned dormancy progression into eight distinct transcriptional modules and identified those exhibiting CR-associated divergence. Candidate-restricted WGCNA resolved these CR-enriched genes into co-expression modules, with one module showing a significant positive correlation with ABA abundance. A cross-cultivar consensus network further nominated hub-centered regulatory candidates. Through integrated evidence prioritization that combined temporal divergence, network topology, cross-cultivar support, and functional annotation, we established a ranked and functionally diverse set of CR-associated candidate regulators, including transcription factors, kinase-related components, hormone-associated genes, and chromatin/RNA factors. Together, these analyses reveal a multi-layer regulatory landscape of CR in which divergence is reflected by stage-dependent remodeling of specific transcriptional and hormone-associated programs rather than a global transcriptome shift. This study provides a structured candidate resource for downstream mechanistic dissection and climate-resilient breeding in peach and related woody perennials.