Integrated Multi-Omics Reveals the Developmental Programs and Metabolic Regulation of the Rare Edible Fungus Buchwaldoboletus xylophilus

Buchwaldoboletus xylophilus is a rare saprotrophic edible mushroom in the Boletaceae and is the second bolete species to have been successfully cultivated artificially, yet its developmental molecular regulation remains unexplored. We integrated RNA-seq transcriptomics and untargeted LC-MS/MS metabolomics across five developmental stages (A–E), applying differential expression, alternative splicing, ceRNA network construction, WGCNA, machine learning, consensus clustering, and O2PLS joint modelling. Transcriptomic analysis revealed that the mycelium-to-primordium transition (stage A to C) showed the most drastic reprogramming, with 2233 differentially expressed genes, while metabolomic comparison between stage A and E revealed overwhelmingly upregulated metabolites (184 up vs. 10 down), indicating asymmetric regulation between the two omics layers. Notably, mutually exclusive exon splicing events exhibited high functional significance, and all top ceRNA hubs were lncRNAs. Unsupervised consensus clustering validated the molecular distinctness of the sampled stages. Metabolite enrichment highlighted antioxidant functions and ABC transporter pathways, and O2PLS integrative modelling confirmed a strong shared developmental signal between transcriptome and metabolome. This study provides the first multidimensional molecular atlas for B. xylophilus, revealing a non-linear developmental programme, transcriptome–metabolome decoupling, and a central role of antioxidant defence throughout development. Our findings offer mechanistic clues regarding the evolutionary transition from ectomycorrhizal to saprotrophic lifestyles in the Boletaceae and establish a multi-omics foundation for the molecular breeding of this emerging edible and medicinal mushroom.