College of Animal Science and Technology, Ningxia University, Yinchuan 750021, China; Key Laboratory of Ruminant Molecular Cell Breeding in Ningxia, Ningxia University, Yinchuan 750021, China.
School of Life Sciences, Yan'an University, Yan'an 716000, China.
Int J Biol Macromol. 2023 Dec 31;253(Pt 4):127096. doi: 10.1016/j.ijbiomac.2023.127096. Epub 2023 Sep 26.
MicroRNAs (miRNAs) are important post-transcriptional factors involved in the regulation of gene expression and play crucial roles in biological processes related to milk fat metabolism. Our previous study revealed that miR-19a expression was significantly higher in the mammary epithelial cells of high-milk fat cows than in those of low-milk fat cows. However, the precise molecular mechanisms underlying these differences remain unclear. In this study, we found a high expression of miR-19a in the mammary tissues of dairy cows. The regulatory effects of miR-19a on bovine mammary epithelial cells (BMECs) were analyzed using cell counting kit-8 and 5-ethynyl-2'-deoxyuridine assays, which demonstrated that miR-19a significantly inhibited BMEC proliferation. Transfection of the miR-19a mimic into BMECs significantly upregulated the expression of milk fat marker genes LPL, SCAP, and SREBP1, promoting triglyceride (TG) synthesis and lipid droplet formation, whereas the miR-19a inhibitor exhibited the opposite function. TargetScan and miRWalk predictions revealed that synaptotagmin 1 (SYT1) is a target gene of miR-19a. A dual luciferase reporter gene assay, RT-qPCR, and western blot analyses revealed that miR-19a directly targets the 3'-untranslated region (UTR) of SYT1 and negatively regulates SYT1 expression. Functional validation revealed that overexpression of SYT1 in BMECs significantly downregulated the expression of LPL, SCAP, and SREBP1, and inhibited TG synthesis and lipid droplet formation. Conversely, the knockdown of SYT1 had the opposite effect. Altogether, miR-19a plays a crucial role in regulating the proliferation and differentiation of BMECs and regulates biological processes related to TG synthesis and lipid droplet formation by suppressing SYT1 expression. These findings provide a strong foundation for further research on the functional mechanisms underlying milk fat metabolism in dairy cows.
微小 RNA（miRNAs）是重要的转录后调控因子，参与基因表达调控，在与乳脂代谢相关的生物学过程中发挥着关键作用。我们之前的研究表明，高乳脂奶牛的乳腺上皮细胞中 miR-19a 的表达明显高于低乳脂奶牛。然而，这些差异的精确分子机制尚不清楚。在这项研究中，我们发现奶牛乳腺组织中 miR-19a 的表达较高。通过细胞计数试剂盒-8 和 5-乙炔基-2'-脱氧尿苷测定分析 miR-19a 对牛乳腺上皮细胞（BMECs）的调控作用，结果表明 miR-19a 显著抑制 BMEC 增殖。转染 miR-19a 模拟物可显著上调乳脂标志物基因 LPL、SCAP 和 SREBP1 的表达，促进甘油三酯（TG）合成和脂滴形成，而 miR-19a 抑制剂则表现出相反的功能。TargetScan 和 miRWalk 预测显示突触结合蛋白 1（SYT1）是 miR-19a 的靶基因。双荧光素酶报告基因检测、RT-qPCR 和 Western blot 分析表明，miR-19a 可直接靶向 SYT1 的 3'-非翻译区（UTR）并负调控 SYT1 的表达。功能验证表明，在 BMECs 中过表达 SYT1 可显著下调 LPL、SCAP 和 SREBP1 的表达，抑制 TG 合成和脂滴形成。相反，SYT1 的敲低则产生相反的效果。综上所述，miR-19a 在调节 BMEC 的增殖和分化中发挥着重要作用，通过抑制 SYT1 的表达，调节与 TG 合成和脂滴形成相关的生物学过程。这些发现为进一步研究奶牛乳脂代谢的功能机制提供了坚实的基础。
