ALKBH5‐mediated RNA m⁶A methylation regulates the migration, invasion and proliferation of rheumatoid fibroblast‐like synoviocytes

Fibroblast-like synoviocytes (FLS) are critical for promoting joint damage in rheumatoid arthritis (RA). N6-methyladenosine (m6 A) modification plays key roles in various diseases; however, its role in the pathogenesis of RA is largely unknown. Here, we investigated increased demethylase ALKBH5 promotes proliferation, migration and invasion of RA FLS via regulating JARID2 expression.ALKBH5 expression in FLS was evaluated using RT-qPCR and WB. EdU, scratch wound healing, and transwell assays were implemented to determine the role of ALKBH5 on RA FLS proliferation, mobility, and migration. Then, m6 A-seq combined with RNA-seq was performed to identify the potential target of ALKBH5. The following RNA immunoprecipitation (RIP) and RNA pulldown was used to validate the interaction between protein and mRNA. CIA and DTHA models were further established to assess the therapeutic potency of ALKBH5 in vivo.We demonstrated that ALKBH5 expression was increased in FLS and synovium from RA. Functionally, ALKBH5 knockdown inhibited the proliferation, migration and invasion of RA FLS, whereas overexpression of ALKBH5 displayed the opposite effect. Mechanistically, ALKBH5 mediated m6 A modification in the JARID2 mRNA and enhanced its mRNA stability in cooperation with IGF2BP3. Intriguingly, the severity of arthritis was attenuated in DTHA mice with ALKBH5 knockout (KO) or CIA rats with intra-articular injection of ALKBH5-shRNA.Our findings suggest that ALKBH5-mediated m6 A modification is crucial for synovial hyperplasia and invasion in RA. ALKBH5 might be a potential therapeutic target for RA and even for dysregulated fibroblasts in a wide range of diseases.