Post-transcriptional effects of leptin 3'UTR polymorphisms on Milk traits in dairy cattle

Leptin plays a central role in regulating energy balance and lactation in dairy cattle. This study aimed to identify single nucleotide polymorphisms (SNPs) in the coding and 3' untranslated regions (3'UTR) of the leptin gene, evaluate their associations with milk production traits, and functionally validate the regulatory effects of key 3'UTR variants on gene expression. Genotyping of 1337 Chinese Holstein cows revealed that c.120 T > C, c.1215 T > C, and c.1761 G > A were polymorphic. The c.120 T > C and c.1215 T > C loci were associated with milk fat, protein, and urea nitrogen levels, whereas c.1761 G > A was associated with increased 305-day milk yield, with cows carrying the GG genotype producing approximately 500 kg more milk than those with the AA genotype. Bioinformatic analyses indicated that the c.1215 T and c.1761 G alleles create potential binding sites for bta-miR-205 and bta-miR-502b, respectively. Dual-luciferase reporter assays confirmed allele-specific miRNA regulation, and Quantitative real-time PCR (qRT-PCR) analysis in mammary tissues and MAC-T cells demonstrated genotype-dependent leptin expression differences. These findings support the hypothesis that 3'UTR polymorphisms modulate leptin expression through miRNA-mediated post-transcriptional regulation. The results provide functional markers, particularly c.1761 G > A, with potential utility for marker-assisted and genomic selection in dairy cattle breeding.