Integrated multi-omics reveals the molecular mechanism of antibiotic-free Wenchang chicken (Gallus Gallus domesticus)

Wenchang chicken is a special local breed in Hainan Province, China, known for its excellent adaptation to the tropical environment and good meat quality. In this study, we compared the differences in serum immunoglobulin levels, antioxidant indices, and liver transcriptome, proteome, phosphoproteomics between antibiotic-free Wenchang chickens (AFS) and normal Wenchang chickens (NS). The IgM level, activities of CAT, TAOC and GSH in serum were significantly higher in AFS Wenchang chickens than in NS Wenchang chickens, but MDA activity in serum was significantly lower in AFS Wenchang chickens. A total of 1,356 DEGs were identified by liver transcriptome analysis and mainly enriched in pathways such as response to bacterium, mitotic cell cycle process, anion binding, and so on. GSEA analysis further indicated that many of the gene sets were associated with immunity regulation and inflammatory responses. A total of 499 DAPs were identified in the liver proteome, mainly involved in the peroxisome, SNARE interactions in vesicular transport and glycerophospholipid metabolism pathways. Three proteins were identified as hubs in the protein interactions network: beta-2-microglobulin precursor (B2M), dedicator of cytokinesis protein 4 (DOCK4) and vesicle-associated membrane protein 3 (VAMP3). There was a weak Spearman's correlation between mRNA expression and protein abundance in the liver. A total of 3,436 phosphoproteins, including 38,891 phosphorylation sites, were identified in the liver of Wenchang chicken. There are 356 phosphoproteins with differential phosphorylation sites, mainly enriched in six pathways, such as the insulin signaling pathway, tight junction and RNA transport pathway. Multi-omics analyses showed that protein phosphorylation in the liver of the Wenchang chicken is largely an independently regulated process. 143 proteins had both up- and down-regulated phosphorylation sites, suggesting that these proteins undergo complex phosphorylation regulation. The AFS Wenchang chickens have higher serum IgM levels and stronger serum antioxidant activity. Accordingly, signaling pathways related to immune and inflammatory responses are highly activated in the livers. The results provide basic data for the disease resistance breeding of Wenchang chicken in the future.