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 <i>leptin</i> 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 <i>leptin</i> expression differences. These findings support the hypothesis that 3'UTR polymorphisms modulate <i>leptin</i> 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.