Alteration of mRNA 5-Methylcytosine Modification in Neurons After OGD/R and Potential Roles in Cell Stress Response and Apoptosis

Abstract

Epigenetic modifications play an important role in central nervous system disorders. As a widespread posttranscriptional RNA modification, the role of the m5C modification in cerebral ischemia-reperfusion injury (IRI) remains poorly defined. Here, we successfully constructed a neuronal oxygen-glucose deprivation/reoxygenation (OGD/R) model and obtained an overview of the transcriptome-wide m5C profiles using RNA-BS-seq. We discovered that the distribution of neuronal m5C modifications was highly conserved, significantly enriched in CG-rich regions and concentrated in the mRNA translation initiation regions. After OGD/R, modification level of m5C increased, whereas the number of methylated mRNA genes decreased. The amount of overlap of m5C sites with the binding sites of most RNA-binding proteins increased significantly, except for that of the RBM3-binding protein. Moreover, hypermethylated genes in neurons were significantly enriched in pathological processes, and the hub hypermethylated genes RPL8 and RPS9 identified by the protein-protein interaction network were significantly related to cerebral injury. Furthermore, the upregulated transcripts with hypermethylated modification were enriched in the processes involved in response to stress and regulation of apoptosis, and these processes were not identified in hypomethylated transcripts. In final, we verified that OGD/R induced neuronal apoptosis in vitro using TUNEL and western blot assays. Our study identified novel m5C mRNAs associated with ischemia-reperfusion in neurons, providing valuable perspectives for future studies on the role of the RNA methylation in cerebral IRI.

Roles of RNA Modifications in Diverse Cellular Functions

“…The role of m 5 C in cell death is not widely explored. Accordingly, one study employing an oxygen-glucose deprivation/reoxygenation (OGD/R) model in neurons found that m 5 C-methylated sites were increased upon OGD/R ( Jian et al, 2021 ). Furthermore, RNA bisulfite sequencing revealed that m 5 C hypermethylated transcripts after OGD/R were functionally involved in apoptosis ( Jian et al, 2021 ).…”

“…Accordingly, one study employing an oxygen-glucose deprivation/reoxygenation (OGD/R) model in neurons found that m 5 C-methylated sites were increased upon OGD/R ( Jian et al, 2021 ). Furthermore, RNA bisulfite sequencing revealed that m 5 C hypermethylated transcripts after OGD/R were functionally involved in apoptosis ( Jian et al, 2021 ). While the functional significance of these findings remains to be explored, these results suggest that m 5 C methylation may be important in mediating apoptosis in response to cellular stress mediated by OGD/R ( Jian et al, 2021 ).…”

“…Furthermore, RNA bisulfite sequencing revealed that m 5 C hypermethylated transcripts after OGD/R were functionally involved in apoptosis ( Jian et al, 2021 ). While the functional significance of these findings remains to be explored, these results suggest that m 5 C methylation may be important in mediating apoptosis in response to cellular stress mediated by OGD/R ( Jian et al, 2021 ). Furthermore, MISU, an NSUN2 homolog, was identified as a critical regulator of mitotic integrity; accordingly, MISU depletion resulted in apoptosis, potentially through decreased spindle integrity and induction of aneuploidy ( Hussain et al, 2009 ).…”

UPLC-MS/MS method for simultaneously determining nucleosides and methyl-nucleosides in liver mRNA of Epimedin C-induced liver injury mouse model

“…For example, m 6 A alteration was associated with the progression and metastasis of hepatocellular carcinoma [ 12 , 13 ], and acetaminophen-induced liver injury in mice [ 14 ]. Besides, m 5 C played an important role in regulation of cell stress response, apoptosis, and hepatocellular carcinoma [ 15 , 16 ]. However, the relationship between mRNA methylation and Epimedin C-induced liver injury is still unknown.…”

Development of A LC-MS/MS Method for Simultaneously Determining Nucleosides and Methy-Nucleosides in Mice Liver mRNA of Epimedin C-Induced Liver Injury Model​

“…Studies indicated that m6A is associated with the progression and metastasis of hepatocellular carcinoma [12][13]. Alteration of m5C in Neurons may lead to cell stress response and apoptosis [14]. The level of m5C modi cation is closely related to PI3K/Akt signaling pathway in gastrointestinal malignancies [15].…”