Analysis of mRNA m6A modification and mRNA expression profiles in middle ear cholesteatoma

Shumin Xie; Li Jin; Jun He; Jinfeng Fu; Tuanfang Yin; Jihao Ren; Wei Liu

doi:10.3389/fgene.2023.1188048

Analysis of mRNA m⁶A modification and mRNA expression profiles in middle ear cholesteatoma

Front Genet. 2023 Aug 7:14:1188048. doi: 10.3389/fgene.2023.1188048. eCollection 2023.

Authors

Shumin Xie¹, Li Jin², Jun He², Jinfeng Fu², Tuanfang Yin², Jihao Ren², Wei Liu²

Affiliations

¹ Hunan Provincial Key Lab, Department of Otolaryngology-Head and Neck Surgery, The Xiangya Hospital, Otolaryngology Institute of Major Diseases, Central South University, Changsha, Hunan, China.
² Department of Otolaryngology-Head and Neck Surgery, The Second Xiangya Hospital, Central South University, Changsha, Hunan, China.

Abstract

Introduction: Middle ear cholesteatoma is characterized by the hyperproliferation of keratinocytes. In recent decades, N⁶-methyladenosine (m⁶A) modification has been shown to play an essential role in the pathogenesis of many proliferative diseases. However, neither the m⁶A modification profile nor its potential role in the pathogenesis of middle ear cholesteatoma has currently been investigated. Therefore, this study aimed to explore m⁶A modification patterns in middle ear cholesteatoma. Materials and methods: An m⁶A mRNA epitranscriptomic microarray analysis was performed to analyze m⁶A modification patterns in middle ear cholesteatoma tissue (n = 5) and normal post-auricular skin samples (n = 5). Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses were performed to predict the potential biological functions and signaling pathways underlying the pathogenesis of middle ear cholesteatoma. Subsequently, m⁶A modification levels were verified by methylated RNA immunoprecipitation-qPCR (MeRIP-qPCR) in middle ear cholesteatoma tissue and normal skin samples, respectively. Results: A total of 6,865 distinctive m⁶A-modified mRNAs were identified, including 4,620 hypermethylated and 2,245 hypomethylated mRNAs, as well as 9,162 differentially expressed mRNAs, including 4,891 upregulated and 4,271 downregulated mRNAs, in the middle ear cholesteatoma group relative to the normal skin group. An association analysis between methylation and gene expression demonstrated that expression of 1,926 hypermethylated mRNAs was upregulated, while expression of 2,187 hypomethylated mRNAs and 38 hypermethylated mRNAs was downregulated. Moreover, GO analysis suggested that differentially methylated mRNAs might influence cellular processes and biological behaviors, such as cell differentiation, biosynthetic processes, regulation of molecular functions, and keratinization. KEGG pathway analysis demonstrated that the hypermethylated transcripts were involved in 26 pathways, including the Hippo signaling pathway, the p53 signaling pathway, and the inflammatory mediator regulation of transient receptor potential (TRP) channels, while the hypomethylated transcripts were involved in 13 pathways, including bacterial invasion of epithelial cells, steroid biosynthesis, and the Hippo signaling pathway. Conclusion: Our study presents m⁶A modification patterns in middle ear cholesteatoma, which may exert regulatory roles in middle ear cholesteatoma. The present study provides directions for mRNA m⁶A modification-based research on the epigenetic etiology and pathogenesis of middle ear cholesteatoma.

Keywords: expression profile; m6A; mRNA; methylation profile; middle ear cholesteatoma.

Grants and funding

This work was supported by the National Natural Science Foundation of China (grant numbers 82071036 and 82000973) and the Natural Science Foundation of Hunan Province (grant numbers 2019JJ50967 and 2022JJ30821).