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Published on: December 23, 2018
Bta-miR-200a Regulates Milk Fat Biosynthesis by Targeting IRS2 to Inhibit the PI3K/Akt Signal Pathway in Bovine
Jianbing Tan1, Benshun Yang1, Liang Qiu1
1College of Animal Science and Technology, Northwest A&F University, Yangling 712100, China.
Abstract:
Milk fat synthesis has garnered significant attention due to its influence on the quality of milk. Recently, an increasing amount of proofs have elucidated that microRNAs (miRNAs) are important post-transcriptional factor involved in regulating gene expression and play a significant role in milk fat synthesis. MiR-200a was differentially expressed in the mammary gland tissue of dairy cows during different lactation periods, which indicated that miR-200a was a candidate miRNA involved in regulating milk fat synthesis. In our research, we investigated the potential function of miR-200a in regulating milk fat biosynthesis in bovine mammary epithelial cells (BMECs). We discovered that miR-200a inhibited cellular triacylglycerol (TAG) synthesis and suppressed lipid droplet formation; at the same time, miR-200a overexpression suppressed the mRNA and protein expression of milk fat metabolism-related genes, such as fatty acid synthase (FASN), peroxisome proliferator-activated receptor gamma (PPARγ), sterol regulatory element-binding protein 1 (SREBP1), CCAAT enhancer binding protein alpha (CEBPα), etc. However, knocking down miR-200a displayed the opposite results. We uncovered that insulin receptor substrate 2 (IRS2) was a candidate target gene of miR-200a through the bioinformatics online program TargetScan. Subsequently, it was confirmed that miR-200a directly targeted the 3'-untranslated region (3'-UTR) of IRS2 via real-time fluorescence quantitative PCR (RT-qPCR), western blot analysis, and dual-luciferase reporter gene assay. Additionally, IRS2 knockdown in BMECs has similar effects to miR-200a overexpression. Our research set up the mechanism by which miR-200a interacted with IRS2 and discovered that miR-200a targeted IRS2 and modulated the activity of the PI3K/Akt signaling pathway, thereby taking part in regulating milk fat synthesis in BMECs. Our research results provided valuable information on the molecular mechanisms for enhancing milk quality from the view of miRNA-mRNA regulatory networks.
Insights
MicroRNAs (miRNAs) regulate milk fat synthesis. MiR-200a inhibits fat synthesis by targeting insulin receptor substrate 2 (IRS2), impacting PI3K/Akt signaling in bovine mammary cells.
Area of Science:
- Molecular Biology
- Animal Science
- Biochemistry
Background:
- Milk fat synthesis is crucial for milk quality and is regulated by complex molecular mechanisms.
- MicroRNAs (miRNAs) are key post-transcriptional regulators of gene expression, with emerging roles in milk fat biosynthesis.
- MiR-200a shows differential expression in dairy cow mammary glands, suggesting its involvement in milk fat regulation.
Purpose of the Study:
- To investigate the function of miR-200a in regulating milk fat biosynthesis in bovine mammary epithelial cells (BMECs).
- To identify the target genes and molecular pathways affected by miR-200a in BMECs.
- To elucidate the regulatory network of miR-200a in milk fat synthesis for potential milk quality enhancement.
Main Methods:
- Overexpression and knockdown of miR-200a in BMECs.
- Measurement of cellular triacylglycerol (TAG) synthesis and lipid droplet formation.
- Analysis of milk fat metabolism-related gene expression (FASN, PPARγ, SREBP1, CEBPα) via RT-qPCR and western blot.
- Bioinformatic prediction and experimental validation (dual-luciferase reporter assay) of miR-200a targets, specifically IRS2.
- Investigation of the PI3K/Akt signaling pathway.
Main Results:
- MiR-200a overexpression inhibited TAG synthesis and lipid droplet formation in BMECs.
- MiR-200a suppressed the expression of key milk fat synthesis genes (FASN, PPARγ, SREBP1, CEBPα).
- Insulin receptor substrate 2 (IRS2) was identified as a direct target of miR-200a.
- MiR-200a regulated milk fat synthesis by targeting IRS2 and modulating the PI3K/Akt signaling pathway.
Conclusions:
- MiR-200a plays an inhibitory role in milk fat synthesis in bovine mammary epithelial cells.
- The mechanism involves direct targeting of IRS2, leading to the modulation of the PI3K/Akt signaling pathway.
- Understanding the miR-200a-IRS2 interaction provides insights into miRNA-mRNA regulatory networks for improving milk quality.
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