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Updated: Mar 2, 2026
A 5-mC Dot Blot Assay Quantifying the DNA Methylation Level of Chondrocyte Dedifferentiation In Vitro
Published on: May 17, 2017
METTL14 deficiency impairs chondrogenic differentiation via m6A-dependent TRAF4 mRNA regulation in Kashin-Beck
Qian Zhang1, Gaowa Naren1, Jiaojiao Zhang1
1School of Public Health, Health Science Center, Xi'an Jiaotong University, Key Laboratory of Environmental and Endemic Diseases of National Health Commission of the People's Republic of China, Xi'an 710061, Shaanxi, PR China.
Objectives:
This study sought to establish the transcriptome-wide m6A methylome landscape in Kashin-Beck disease (KBD) chondrocytes and delineate the mechanistic contributions of the m6A methyltransferase METTL14 to KBD pathogenesis.
Methods:
Articular cartilage specimens were obtained from surgically confirmed Kashin-Beck disease (KBD) patients and normal controls. Multi-omics integration combined MeRIP-seq and RNA-seq to delineate m6A landscapes, supplemented by spectrophotometric m6A quantification. Differential expression of m6A machinery components was validated using qRT-PCR and immunoblotting. In chondrocyte models, lentiviral-mediated METTL14 knockdown (shRNA) and complementary overexpression were performed. METTL14-TRAF4 interaction was resolved via MeRIP and FISH. The therapeutic mechanism of quercetin was analyzed using CETSA and molecular docking.
Results:
The methylome profiling of KBD articular cartilage revealed 1539 transcripts with differential m6A methylation patterns. Pathway enrichment analysis identified significant association with Wnt signaling and ubiquitin-mediated proteolysis pathways. Global m6A hypomethylation was accompanied by reduced expression of core methyltransferase complex components, including METTL14, YTHDC2, and WTAP. Mechanistic studies demonstrated that METTL14 directly binds to TRAF4 mRNA through an RNA-protein interaction. METTL14 knockdown significantly attenuated chondrogenic differentiation capacity, as evidenced by reduced expression of COL2A1, SOX9, TGF-β1, and PTHrP, while pharmacological activation via quercetin restored METTL14 expression and ameliorated KBD phenotypes.
Conclusion:
METTL14 is a central epigenetic modulator in KBD pathophysiology through TRAF4 coupled regulation of chondrocyte differentiation.
Insights
This study reveals that METTL14, an epigenetic regulator, is crucial in Kashin-Beck disease (KBD) pathogenesis by influencing chondrocyte differentiation via TRAF4. Quercetin shows therapeutic potential by restoring METTL14 levels.
Area of Science:
- Epigenetics and RNA modification
- Molecular biology of cartilage
- Disease mechanisms
Background:
- Kashin-Beck disease (KBD) is a severe osteochondropathy.
- The role of RNA methylation in KBD pathogenesis is largely unknown.
- m6A modification is critical for cellular processes.
Purpose of the Study:
- To map the m6A methylome in KBD chondrocytes.
- To investigate the role of METTL14 in KBD.
- To explore quercetin's therapeutic potential in KBD.
Main Methods:
- Integrated multi-omics (MeRIP-seq, RNA-seq) on KBD patient cartilage.
- Validation of gene expression via qRT-PCR and immunoblotting.
- Functional studies using chondrocyte models, METTL14 manipulation, and quercetin treatment.
Main Results:
- Identified 1539 differentially methylated transcripts in KBD, linked to Wnt signaling and proteolysis.
- Observed global hypomethylation and reduced METTL14 expression in KBD chondrocytes.
- Demonstrated METTL14 binds TRAF4 mRNA, and its knockdown impairs chondrogenesis; quercetin ameliorates KBD phenotypes.
Conclusions:
- METTL14 is a key epigenetic regulator in KBD pathophysiology.
- METTL14 regulates chondrocyte differentiation through TRAF4.
- Quercetin can restore METTL14 expression and mitigate KBD phenotypes.
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