Hypoxia-induced m6A epitranscriptomic remodeling regulates gene expression in dental pulp stem cells: a combined MeRIP-seq and RNA-seq study

Abstract

Dental pulp stem cells (DPSCs) are essential for regenerative endodontics. However, their survival and function are often challenged by the physiological hypoxic environment of the dental pulp. While N6-methyladenosine (m6A) modification is a critical post-transcriptional regulator of the hypoxic response, its specific landscape and role in DPSCs remain unknown. This study aims to map the m6A epitranscriptomic landscape of DPSCs under hypoxia to identify novel regulatory mechanisms. Human DPSCs were cultured under normoxia and hypoxia. m6A methylated RNA immunoprecipitation sequencing (MeRIP-seq) and RNA sequencing (RNA-seq) were performed to profile m6A modification and gene expression levels, respectively. Bioinformatic analysis identified differentially methylated and expressed genes, which were validated via qRT-PCR and predictive binding analysis. MeRIP-seq revealed that while global m6A abundance in DPSCs remained relatively stable under hypoxia, the topological distribution underwent significant remodeling. Conjoint analysis identified 1,169 differentially methylated peaks (DMPs) with significant changes in methylation alongside expression alterations in their corresponding host genes. Functional enrichment showed these genes are primarily involved in the PI3K-Akt signaling pathway, angiogenesis, and the response to hypoxia. Specifically, the hub genes FAM13A, COL4A1, PPFIA4, and PTGS2 were hyper-methylated and up-regulated. Predictive analysis suggested that the putative reader IGF2BP1 potentially interacts with the m6A sites of these target transcripts. Notably, the involvement of these specific hub genes has not been previously investigated in DPSCs. This study reveals that hypoxia induces targeted m6A epitranscriptomic remodeling in DPSCs. We propose a putative regulatory axis where hypoxia-upregulated IGF2BP1 may facilitate cellular adaptation by stabilizing transcripts associated with angiogenesis and tissue regeneration. Our findings suggest these genes may serve as pivotal regulators of the hypoxic response in DPSCs, representing a novel mechanism for adaptation. While these results provide new insights, further functional validation is required to fully elucidate the specific mechanisms of the IGF2BP1–m6A axis in pulp regeneration.