Identification of Key Genes Regulating Skeletal Development of the Tibia in Chickens Through the Integration of Genome-Wide Association Analysis and Transcriptome Sequencing

Body measurement traits (particularly tibia length and width) are important indicators of poultry growth and development and are key traits that influence production performance and economic value. Since the development of long bones, such as the tibia, directly affects skeletal structure, body size, and growth potential, elucidating their genetic regulatory mechanisms is of great significance for improving poultry production performance and advancing molecular breeding. To elucidate the genetic basis of body measurement traits in TC-line Yellow-Feathered Broiler chickens, this study conducted phenotypic correlation analyses and estimated genetic parameters for individuals across four generations (16–19). The results revealed that tibia length and tibia width were significantly and positively correlated with major slaughter traits (slaughter weight, semi-eviscerated weight, fully eviscerated weight, breast muscle weight, thigh weight, and wing weight). The heritabilities of tibia length and tibia width were 0.52 and 0.74, respectively, indicating that these are traits with high heritability. A genome-wide association study (GWAS) was conducted on 436 samples, identifying 61 and 130 suggestively associated SNP loci for tibia length and tibia width, respectively, which were annotated to 20 and 40 candidate genes. KEGG enrichment analysis of the GWAS candidate genes revealed enrichment in metabolic pathways, the Wnt signaling pathway, and amino acid biosynthesis, among others. Transcriptomic sequencing of the growth plate revealed a total of 522 differentially expressed genes (DEGs) between the high-development (H) and low-development (L) groups, with five roosters per group, of which 296 were upregulated and 226 were downregulated. Tibia length differed significantly between the H and L groups, whereas tibia width did not. Thus, the RNA-seq comparison primarily supports interpretations related to tibia length, and conclusions regarding tibia width should be treated cautiously. Functional enrichment analysis of the DEGs indicated that they were primarily involved in ion binding, cell differentiation, and transcriptional regulation processes related to skeletal development, with enrichment of pathways including FoxO, mTOR, cell adhesion, and lysosomes. Integration of the GWAS and transcriptomic results identified TENM2 as the only overlapping candidate gene for tibia length; no overlapping candidate gene was detected for tibia width. All 18 suggestively associated loci on chromosome 13 (including positions 4,921,436, 4,967,805, and 4,988,787) were annotated to TENM2, which was also differentially expressed between the H and L groups, providing convergent evidence at the genomic and transcriptomic levels. TENM2 (teneurin transmembrane protein 2) encodes a protein whose Gene Ontology (GO) annotations include calcium ion binding and protein heterodimerization activity. This study provides a reference for the genetic improvement of body measurement traits in TC-line Yellow-Feathered Broiler chickens.