Comprehensive Profiling of Transcriptome and m6A Epitranscriptome Uncovers the Neurotoxic Effects of Yunaconitine on HT22 Cells

Abstract

Objective: To explore different mRNA transcriptome patterns and RNA N6-methyladenosine (m6A) alteration in yunaconitine (YA)-treated HT22 mouse hippocampal neuron, and uncover the role of abnormal mRNA expression and RNA m6A modification in YA-induced neurotoxicity. Methods: HT22 cells were treated with 0, 5, 10, and 50 μM of YA for 72 h to evaluate their viability and GSH content. Subsequently, mRNA-seq and MeRIP-seq analyses were performed on HT22 cells treated with 0 and 10 μM YA for 72 h, and molecular docking was used to simulate interactions between YA and differentially expressed m6A regulators. The mitochondrial membrane potential was examined using the JC-10 probe, and RT-qPCR was conducted to verify the expression levels of differentially expressed m6A regulatory factors, as well as to assess alterations in the mRNA expression levels of antioxidant genes. Results: YA treatment significantly reduced the viability of HT22 cells and decreased GSH content. The mRNA-seq analysis obtained 1018 differentially expressed genes, KEGG and GO enrichment results of differentially expressed genes mainly comprise the nervous system development, cholinergic synapse, response to oxidative stress, and mitochondrial inner membrane. A total of 7 differentially expressed m6A regulators were identified by MeRIP-seq. Notably, molecular docking results suggested a stable interaction between YA and most of the differentially expressed m6A regulators. Conclusion: This study showed that YA-induced HT22 cell damage was associated with the increased methylation modification level of target gene m6A and abnormal expression of m6A regulators.