A Functional 3′ UTR Polymorphism of FADS2 Affects Cow Milk Composition through Modifying Mir-744 Binding

SIMPLE SUMMARY: Fatty acid desaturase 2 (FADS2) is the rate-limiting enzyme involved in the synthesis of long-chain polyunsaturated fatty acids (LC-PUFAs). Many studies have suggested that polymorphisms in the FADS2 gene can modify its delta-6 desaturase activity. However, much remains unknown in regards to the regulatory mechanisms interpreting how DNA variants influence the function of FADS2. A previous study has suggested that c.1571G>A is located within the miR-744 binding site, indicating that this single nucleotide polymorphism (SNP) may be functional. Therefore, the aim of the present study was to determine the association of SNP c.1571G>A with milk PUFAs content and to further elucidate how this SNP contributes to regulating FADS2 expression. Our results support the use of SNP c.1571G>A as a potential genetic marker in the selective breeding of cattle to increase beneficial FAs content in milk. ABSTRACT: This study determined the associations of FADS2 c.1571G>A with milk FAs content and revealed that cows with the GG genotype had improved levels of delta-6 desaturase substrates (linoleic acid, C18:2n-6; p < 0.001) and decreased levels of desaturase products (gamma-linolenic acid, C18:3n-6; p < 0.001), indicating a reduction in FADS2 expression or delta-6 desaturase activity caused by this polymorphism. Computer alignment demonstrated that c.1571G>A occurred within a potential miR-744 binding site. When the c.1571G allele was present, the luciferase activity of reporter constructs was significantly suppressed by miR-744, while no such effect was observed with the A allele. Overexpression of miR-744 in bovine mammary epithelial cells (with the 1571GG genotype) downregulated FADS2 expression at both mRNA and protein levels. In contrast, inhibition of endogenous miR-744 with a specific inhibitor dramatically upregulated FADS2 expression. Taken together, these lines of evidence indicated that the c.1571A minor allele abolished the ability of miR-744 to bind FADS2, with a consequent increase in FADS2 expression levels and synthesis of omega-6 LC-PUFAs.