Gene regulatory and co-expression networks reveal novel hub genes and regulatory mechanisms in rapeseed (Brassica napus L.) under drought stress

Rapeseed (Brassica napus L.) productivity is increasingly threatened by drought, yet coordinated regulatory programs that govern drought responses remain incompletely characterized. Here, we integrated an RNA-Seq meta-analysis with weighted gene co-expression network analysis (WGCNA) and gene regulatory network (GRN) inference to study in depth the molecular responses of rapeseed to drought stress. WGCNA revealed three modules: brown module enriched for upregulated genes and two primarily downregulated modules (black and cyan). Brown-module hubs coordinate ABA signaling, stomatal regulation, lipid mobilization, cell-wall reinforcement and protein homeostasis, whereas black and cyan module hubs converge on suppression of growth, defense and metal-detoxification pathways and reallocation of resources away from reproduction. Furthermore, this study identified crucial hub transcription factors (TFs) within constructed GRNs. Upregulated TFs orchestrated the induction of downstream genes involved in ABA signaling, protein homeostasis, antioxidant and redox maintenance, stomatal, calcium, and ion regulation, membrane, cell wall, and lipid remodeling, photosynthetic processes, and developmental reprogramming. Conversely, TFs in the downregulated GRN repress growth-promoting signals, stomatal opening, and defense responses, while concurrently regulating water use efficiency, water transport, and nutrient uptake. Our results suggest that the ABA-mediated drought response in rapeseed involves an intricate interplay of positive and negative regulators that prevents excessive ABA activation; concurrently, protein homeostasis is maintained by prioritizing repair and refolding over wholesale degradation. Several TFs and hub genes identified in this study, including BHLH122-1, HHO6, RAB18, LTP4, LEA30, DAA1, and EXLB1, represent promising candidates for genomic selection and biotechnology approaches aimed at improving drought resilience in rapeseed.