Dynamic changes in genes and metabolites during embryonic development of Jiangshan black-bone chickens

Melanin deposition in the breast muscle is a defining economic and nutritional trait of Jiangshan black-bone chickens, yet the biological architecture governing this phenotype is intricate. While the phenotypic characteristics are well-documented, the dynamic interplay between transcriptomic regulation and metabolic shifts driving melanin accumulation during critical embryonic stages remains underexplored. This study aimed to elucidate the temporal molecular mechanisms of melanin deposition in breast muscle during embryonic development (Embryonic day 13 - Embryonic day 21, E13-E21) by integrating transcriptome and metabolome analyses. We quantified melanin content and performed RNA-sequencing and LC-MS/MS metabolomics on breast muscle tissue collected across five developmental stages (E13, E15, E17, E19, and E21). Results showed a significant increase in melanin content from E13 to E19, stabilizing by E21. The multi-omics integration identified 8,651 differentially expressed genes (DEGs) and 712 differential metabolites (DMs). Pathway analysis revealed significant enrichment in broader amino acid metabolism, MAPK signaling, and neuroactive ligand-receptor interactions. Furthermore, targeted candidate analysis on the classical melanogenesis network successfully highlighted key regulatory genes such as DCT, TYRP1, and MC1R. Metabolome analyses shows that amino acid metabolism plays an important role in melanin synthesis. Correlation analysis unveiled a tight linkage between metabolites (phenylpyruvate, glycerol, 1-myristoyl-2-palmitoyl-sn‑glycero-3-phosphocholine (PC), 1,2-dipalmitoyl-sn‑glycero-3-phosphoethanolamine (PEA)) and functional genes related to immunity (IL4I1, MLKL) and lipid remodeling (PNPLA2, CCNB1). Furthermore, specific alterations in sulfur metabolism genes (CBS, CTH, GPX1) suggested a regulatory mechanism balancing eumelanin and pheomelanin synthesis. Crucially, the reliability of these transcriptomic and conjoint analyses was robustly confirmed via targeted qRT-PCR validation of key genes regulating melanogenesis, membrane dynamics, and sulfur amino acid metabolism. These findings highlight the intricate interplay between the transcriptome and metabolome underlying muscle hyperpigmentation and provide novel insights into the molecular breeding targets for optimizing meat quality in poultry.