Study on the Suppression of Deubiquitinase Rpn11 to Improve Non-Alcoholic Fatty Liver Disease (NAFLD)

Background Introduction

Non-alcoholic fatty liver disease (NAFLD) and its more severe form, non-alcoholic steatohepatitis (NASH), have become major global public health issues. The rapid increase in their incidence has made them leading causes of cirrhosis and hepatocellular carcinoma (HCC). NAFLD is not only associated with hepatic fat deposition but is also often accompanied by metabolic disorders such as insulin resistance, elevated blood sugar, and hyperlipidemia. In recent years, experts have redefined NAFLD as metabolic dysfunction-associated steatotic liver disease (MASLD), further emphasizing its complex metabolic characteristics.

Currently, the main pathological feature of NAFLD is the excessive accumulation of triglycerides within hepatocytes, a process regulated by multiple transcription factors. However, the impact of post-translational modifications (such as ubiquitination and deubiquitination) on hepatic lipid metabolism remains unclear. Ubiquitin is a conserved protein that can promote protein degradation, signal transduction, and gene transcription by attaching to lysine residues on target proteins. This process is reversible, with deubiquitinases (DUBs) removing ubiquitin to stabilize the expression of target proteins. Research on DUBs in both healthy and disease states has been gradually deepening, but their potential in NAFLD has not yet been fully explored.

In this study, a research team using mice as a model identified Rpn11 as a significantly upregulated DUB in NAFLD mouse models through gene expression screening. Through a series of experiments, they revealed Rpn11’s key role in regulating liver lipid metabolism. The study results indicate that Rpn11 deubiquitinates and stabilizes METTL3, thereby enhancing the m6A modification of ACSS3. This Rpn11-METTL3-ACSS3 axis further upregulates genes related to lipid metabolism through histone propionylation. This new finding suggests that inhibiting Rpn11 could be a novel strategy for treating NAFLD.

Study Source

This study was conducted collaboratively by researchers from Fudan University, Shanghai Jiao Tong University School of Medicine, Wenzhou Medical University, and Shanghai Institute of Materia Medica. Authors include Bing Zhou, Yunchen Luo, Hanqi Bi, among others. The article was published on October 1, 2024, in the journal “Cell Metabolism.”

Research Process

The research team employed a multi-step experimental design to investigate the specific mechanism of Rpn11 in NAFLD and NASH.

1. Model Establishment: To study the role of Rpn11, researchers constructed hepatocyte-specific Rpn11 knockout mice (Rpn11 HKO) to observe its performance in NAFLD-related phenotypes. Simultaneously, the team induced NAFLD and NASH in mice through high-fat diet (HFD) and Gubra Amy NASH (GAN) diet methods. Additionally, they used leptin receptor-deficient mice (db/db) and employed qPCR, RNA sequencing, and western blotting to detect Rpn11 expression in various mouse NAFLD models.

2. Relationship Between Rpn11 and METTL3: The study found that Rpn11 stabilizes the METTL3 protein through deubiquitination, thus upregulating its expression levels. Specific experiments showed that in Rpn11 HKO mice, METTL3 protein significantly decreased, although there was no apparent change in its mRNA levels. Further study showed that Rpn11 can increase the stability of METTL3 protein in HFD-induced steatohepatitis mice.

3. METTL3 Regulates ACSS3 Expression: Rpn11-stabilized METTL3 promotes ACSS3 expression through m6A modification. ACSS3 is a short-chain acyl-CoA synthetase that can generate propionyl-CoA, subsequently upregulating the expression of lipid metabolism-related genes through histone propionylation.

4. Efficacy Experiment on Inhibiting Rpn11: The research team used the Rpn11 inhibitor Capzimin to treat db/db mice, finding that the inhibitor significantly improved liver steatosis, insulin resistance, and blood sugar levels in the mice. Capzimin not only reduced METTL3 protein levels but also decreased ACSS3 expression, histone propionylation levels, and the expression of lipid metabolism-related genes.

Main Research Results

1. Key Role of Rpn11 Upregulation in NAFLD: Rpn11 is significantly upregulated in HFD and GAN diet-induced NAFLD/NASH mouse models, and its impact on hepatic lipid accumulation was observed in liver-specific Rpn11 knockout mice. Results indicated that Rpn11 HKO mice have significant protective effects against high-fat diet-induced liver steatosis.

2. Discovery of the Rpn11-METTL3-ACSS3 Pathway: Rpn11 deubiquitinates and stabilizes METTL3, which in turn enhances the m6A modification level of ACSS3, increasing ACSS3 protein expression. Propionyl-CoA produced by ACSS3 regulates the expression of lipid metabolism genes through histone propionylation, promoting the development of NAFLD.

3. Therapeutic Potential of Capzimin: The Rpn11 inhibitor Capzimin demonstrated good therapeutic effects in the db/db mouse model, significantly reducing hepatic lipid accumulation and improving insulin sensitivity and glucose tolerance in mice. This suggests Capzimin may become a new drug candidate for treating NAFLD and NASH.

Conclusion and Significance

This study reveals the important role of Rpn11 as a novel regulatory factor in NAFLD and NASH. Through deubiquitination, Rpn11 stabilizes METTL3, thereby promoting ACSS3 m6A modification and activating lipid metabolism genes. The discovery of this Rpn11-METTL3-ACSS3-histone propionylation axis not only enriches understanding of the pathogenic mechanisms of NAFLD but also provides new ideas for the treatment of this disease.

This study opens new avenues for potential therapeutic options for NAFLD, proposing that inhibiting Rpn11 might become an effective treatment strategy. This strategy holds especially important application prospects for patients with NAFLD and NASH. In the future, drug development such as with Capzimin may provide patients with more effective treatment options. Furthermore, since METTL3 plays a key role in various physiological and pathological processes, research on the Rpn11-METTL3 pathway may also have important implications for other diseases, such as cancer and immune-related diseases.

Research Highlights

1. Discovery of a New Regulatory Pathway: The study innovatively reveals the critical influence of Rpn11 on liver lipid metabolism via the Rpn11-METTL3-ACSS3 pathway in NAFLD development.

2. Exploration of Therapeutic Potential: The successful application of Capzimin in mouse models demonstrates its great potential in NAFLD and NASH treatment, offering a new direction for clinical therapy in this field.

3. In-depth Understanding of NAFLD Pathological Mechanisms: Through various experimental models and techniques, the research team elucidated the mechanism of Rpn11’s role in NAFLD, providing a solid theoretical foundation for subsequent research.

Outlook

Future research can further explore the impact of Rpn11 on other metabolic diseases beyond NAFLD, particularly the potential role of the Rpn11-METTL3 pathway in diseases such as tumors. Moreover, the safety and tolerance of Capzimin need further validation in clinical trials, especially concerning its therapeutic effects in female patients. This study provides new ideas for the treatment of NAFLD/NASH, carrying significant scientific and applied value.