Related Experiment Video
Updated: Jun 30, 2025
A Reporter Assay to Analyze Intronic microRNA Maturation in Mammalian Cells
Published on: June 16, 2022
Excessive fatty acids activate PRMT5/MDM2/Drosha pathway to regulate miRNA biogenesis and lipid metabolism
Aijun Hou1,2, Xiaoding Xu1,3, Yu Zhang1,2
1Center for Drug Safety Evaluation and Research, State Key Laboratory of Drug Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai, China.
Background:
Excessive fatty acids in the liver lead to the accumulation of lipotoxic lipids and then cellular stress to further evoke the related disease, like non-alcoholic fatty liver disease (NAFLD). As reported, fatty acid stimulation can cause some specific miRNA dysregulation, which caused us to investigate the relationship between miRNA biogenesis and fatty acid overload.
Methods:
Gene expression omnibus (GEO) dataset analysis, miRNA-seq, miRNA cleavage assay, RT-qPCR, western blotting, immunofluorescence and co-immunoprecipitation (co-IP) were used to reveal the change of miRNAs under pathological status and explore the relevant mechanism. High fat, high fructose, high cholesterol (HFHFrHC) diet-fed mice transfected with AAV2/8-shDrosha or AAV2/8-shPRMT5 were established to investigate the in vivo effects of Drosha or PRMT5 on NAFLD phenotype.
Results:
We discovered that the cleavage of miRNAs was inhibited by analysing miRNA contents and detecting some representative pri-miRNAs in multiple mouse and cell models, which was further verified by the reduction of the Microprocessor activity in the presence of palmitic acid (PA). In vitro, PA could induce Drosha, the core RNase III in the Microprocessor complex, degrading through the proteasome-mediated pathway, while in vivo, knockdown of Drosha significantly promoted NAFLD to develop to a more serious stage. Mechanistically, our results demonstrated that PA can increase the methyltransferase activity of PRMT5 to degrade Drosha through MDM2, a ubiquitin E3 ligase for Drosha. The above results indicated that PRMT5 may be a critical regulator in lipid metabolism during NAFLD, which was confirmed by the knocking down of PRMT5 improved aberrant lipid metabolism in vitro and in vivo.
Conclusions:
We first demonstrated the relationship between miRNA dosage and NAFLD and proved that PA can activate the PRMT5-MDM2-Drosha signalling pathway to regulate miRNA biogenesis.
Insights
Excessive fatty acids in non-alcoholic fatty liver disease (NAFLD) disrupt microRNA (miRNA) biogenesis. Palmitic acid activates PRMT5-MDM2-Drosha signaling, impairing miRNA production and worsening NAFLD progression.
Area of Science:
- Molecular Biology
- Hepatology
- Biochemistry
Background:
- Excessive hepatic fatty acids cause lipotoxicity and cellular stress, contributing to non-alcoholic fatty liver disease (NAFLD).
- Fatty acid overload is known to dysregulate specific microRNAs (miRNAs), prompting investigation into miRNA biogenesis.
- Understanding the link between fatty acids and miRNA regulation is crucial for NAFLD pathogenesis.
Purpose of the Study:
- To investigate the relationship between fatty acid overload and miRNA biogenesis in the context of NAFLD.
- To elucidate the molecular mechanisms by which fatty acids affect miRNA production.
- To identify key regulators involved in fatty acid-induced miRNA dysregulation in NAFLD.
Main Methods:
- Analysis of Gene Expression Omnibus (GEO) datasets and miRNA sequencing (miRNA-seq).
- In vitro assays including miRNA cleavage assay, RT-qPCR, western blotting, immunofluorescence, and co-immunoprecipitation (co-IP).
- In vivo studies using high-fat, high-fructose, high-cholesterol diet-fed mice with AAV2/8-mediated knockdown of Drosha or PRMT5.
Main Results:
- Palmitic acid (PA) inhibited miRNA cleavage and reduced Microprocessor activity, indicating impaired miRNA biogenesis.
- PA induced the proteasomal degradation of Drosha, a key miRNA processing enzyme, exacerbating NAFLD phenotype in vivo.
- PA increased PRMT5 methyltransferase activity, leading to Drosha degradation via MDM2, a ubiquitin E3 ligase. Knockdown of PRMT5 improved lipid metabolism.
Conclusions:
- Established a direct link between miRNA dosage and NAFLD development.
- Demonstrated that palmitic acid activates the PRMT5-MDM2-Drosha signaling pathway, regulating miRNA biogenesis.
- Identified PRMT5 as a critical regulator of lipid metabolism in NAFLD, with its inhibition ameliorating the disease phenotype.
Related Concept Videos
01:22MicroRNAs
01:23Regulation of Expression at Multiple Steps
02:51mRNA Stability and Gene Expression
Cis-acting Elements involved in mRNA stability
02:24Regulation of Expression Occurs at Multiple Steps
Transcription results in the generation of precursor (pre-mRNA) that consists of both exons and introns, which needs further processing before being translated to a...
02:31Nuclear Export of mRNA
01:24Overview of Lipid Metabolism
Lipolysis: The Breakdown of Lipids:
Lipolysis is the process of breaking down lipids, particularly triglycerides, into glycerol and fatty acids. This process typically occurs in the adipose tissue and is triggered by various hormones, including glucagon and...