Elevated N6-Methyladenosine (m6A)-RNA-Methylation During Oral Carcinogenesis.

Background/purpose: Oral epithelial dysplasia (OED) and oral squamous cell carcinoma (OSCC) encompass a series of molecular events in the malignant transformation process, ranging from simple epithelial hyperplasia to mild, moderate, and severe dysplasia. N6-methyladenosine (m6A)-RNA methylation participates in the regulation of the tumorigenesis of various malignant tumors, yet the roles played by m6A-RNA methylation in OED and OSCC remain unclear. Therefore, this study focused on investigating OED and OSCC from an epigenetic perspective, aiming to elucidate the underlying molecular mechanisms of malignant transformation.

Materials and methods: Laser microdissection was performed on OED and OSCC samples. Methylated RNA immunoprecipitation sequencing (MeRIP-Seq) and RNA sequencing (RNA-seq) were applied to establish comprehensive profiles of m6A methylation modifications and gene expression patterns and to identify differentially modified/expressed genes in OED and OSCC.

Results: We presented the overall modification/expression profiles of m6A-RNA-methylation in OED and OSCC. Four hypermethylated genes and 11 hypomethylated genes were found in both OED and OSCC, together with the expression of 107 upregulated and 37 downregulated genes. The most common motifs GRAGRA (R = A/G) of the OED and OSCC methylation sites were mainly located in coding and stop codon regions. In the stable group, C4B, DNAH9, and NCALD all exhibited hypermethylated and upregulated, and the overall survival rate of patients with high expression of these genes was higher than that of patients with low-level expression.

Conclusion: Our study revealed that the level of m6A-RNA methylation in the epithelial tissues of OED and OSCC was higher than that in oral normal epithelium, suggesting that the methylation modification might be involved in the occurrence of OED and its progression to OSCC. Furthermore, hypermethylation and upregulated expression of C4B, DNAH9, and NCALD were associated with a favorable prognosis in these diseases.