Comprehensive Analysis and Functional Characteristics of Differential Expression of N6-Methyladenosine Methylation Modification in the Whole Transcriptome of Rheumatoid Arthritis

Abstract

N6-methyladenosine (m6A) modification is the most prevalent chemical modification in eukaryotic mRNA and is associated with the development of various immune diseases. However, the role of m6A methylation in rheumatoid arthritis (RA) development is unclear. We preliminarily explored the role of m6A methylation-related mRNAs in RA for its clinical application. The discovery of m6A methylation-modifying genes in this study may provide a fresh perspective on the development of drugs for RA treatment. High-throughput sequencing combined with methylated RNA immunoprecipitation (MeRIP-seq) and RNA sequencing were used to assess whole-transcriptome m6A modifications in the synovium of patients with RA. The relationship between m6A-modified target genes and RA inflammation and macrophages was determined. The expression of the m6A-modified significant transcript-enriched inflammatory signaling pathway was assessed through animal experiments. Differentially expressed m6A genes were correlated with macrophage activation involved in immune response, vascular endothelium, MAPK signaling pathway, PI3K - Akt signaling pathway, and other inflammatory processes. Furthermore, combined analysis with m6A-seq and RNA-seq revealed 120 genes with significant changes in both m6A modification and mRNA expression. We selected the top 3 candidate mRNAs that were upregulated and downregulated simultaneously. The expression of phosphatase and tensin homolog deleted on chromosome ten (PTEN) mRNA and protein in RA patients was lower than that in healthy control (HC). SHC-binding protein 1 (SHCBP1) and neurexophilin-3 (NXPH3) mRNA expressions were increased in RA patients. The expression of M1 macrophages was increased in RA patients. RA markers are such as rheumatoid factor (RF) and peptide containing citrulline (CCP). Further animal experiments showed that the expression of synovial MAPK, PI3K, and Akt1 proteins in the RA model was increased, and the PTEN, p-PTEN protein expression was decreased. PI3K, Akt1, PTEN, and p-PTEN were correlated to RA joint inflammation. This study revealed a unique pattern of differential m6A methylation modifications in RA and concluded that m6A modification is related to the occurrence of RA synovial inflammation.