Shaanxi Key Laboratory of Molecular Biology for Agriculture, College of Animal Science and Technology, Northwest A&F University, Yangling, Xianyang Shaanxi 712100, PR China.
College of Life Science and Agricultural Engineering, Nanyang Normal University, Nanyang, Henan 473061, PR China.
J Agric Food Chem. 2023 May 17;71(19):7440-7450. doi: 10.1021/acs.jafc.2c09001. Epub 2023 May 8.
MicroRNA-mediated gene regulation is important for the regulation of fatty acid metabolism and synthesis. Our previous study uncovered that the miR-145 expression is higher in the lactating mammary gland of dairy cows than in the dry-period, but the underlying molecular mechanism is incompletely understood. In this study, we have investigated the potential role of miR-145 in bovine mammary epithelial cells (BMECs). We found that the expression of miR-145 gradually increased during lactation. CRISPR/Cas9-mediated knockout (KO) of miR-145 in BMECs results in the downregulated expression of fatty acid metabolism-associated genes. Further results revealed that miR-145 KO reduced total triacylglycerol (TAG) and cholesterol (TC) accumulation and altered the composition of intracellular fatty acids (C16:0, C18:0, and C18:1). Conversely, miR-145 overexpression had the opposite effect. Bioinformatics online program predicted that miR-145 targets the 3'-UTR of the Forkhead Box O1 () gene. Subsequently, was identified as a direct target of miR-145 by qRT-PCR, Western blot analysis, and luciferase reporter assay. Furthermore, siRNA-mediated silencing of promoted fatty acid metabolism and TAG synthesis in BMECs. Additionally, we observed the involvement of in the transcriptional activity of the sterol regulatory element-binding protein 1 () gene promoter. Overall, our findings indicated that miR-145 relieves the inhibitory effect of on expression by targeting and subsequently regulating fatty acid metabolism. Thus, our results provide valuable information on the molecular mechanisms for improving milk yield and quality from the perspective of miRNA-mRNA networks.
miRNA 介导的基因调控对于脂肪酸代谢和合成的调节至关重要。我们之前的研究表明，奶牛泌乳期乳腺中的 miR-145 表达高于干奶期，但其中的分子机制尚不完全清楚。在这项研究中，我们研究了 miR-145 在牛乳腺上皮细胞（BMEC）中的潜在作用。我们发现，miR-145 的表达在泌乳期逐渐增加。CRISPR/Cas9 介导的 miR-145 在 BMEC 中的敲除（KO）导致脂肪酸代谢相关基因的下调表达。进一步的结果表明，miR-145 KO 减少了总三酰甘油（TAG）和胆固醇（TC）的积累，并改变了细胞内脂肪酸（C16:0、C18:0 和 C18:1）的组成。相反，miR-145 的过表达则有相反的效果。生物信息学在线程序预测 miR-145 靶向 Forkhead Box O1 () 基因的 3'-UTR。随后，通过 qRT-PCR、Western blot 分析和荧光素酶报告基因测定，确定 是 miR-145 的直接靶标。此外，siRNA 介导的 沉默促进了 BMEC 中的脂肪酸代谢和 TAG 合成。此外，我们观察到 参与了固醇调节元件结合蛋白 1 () 基因启动子的转录活性。总的来说，我们的研究结果表明，miR-145 通过靶向 并随后调节脂肪酸代谢来解除 对 表达的抑制作用。因此，我们的研究结果从 miRNA-mRNA 网络的角度为提高牛奶产量和质量提供了有价值的信息。
