Neuronal Depolarization Induced RNA m(5)C Methylation Changes in Mouse Cortical Neurons

SIMPLE SUMMARY: When neurons are activated, two waves of gene expression changes occur to achieve neuronal adaptation to their environment. Currently, little is known about the changes in post-transcriptional modifications of neuronal transcripts in response to environmental stimuli. This study aims to investigate the dynamics of m(5)C modification on neuronal mRNAs upon depolarization using potassium chloride. We found that neuronal mRNAs carrying m(5)C modifications are enriched for genes with important neuronal functions. The numbers of m(5)C sites and methylated transcripts increased at 2 h after neuronal depolarization and slightly dropped at 6 h. Interestingly, for differentially methylated genes, a negative correlation between the levels of gene expression and RNA methylation is more prevalent. We anticipate our findings may shed new lights on post-transcriptional regulation in activated neurons, and the transcriptomic plus epi-transcriptomic datasets generated in this study would provide a valuable resource to the scientific community. ABSTRACT: Neuronal activity is accomplished via substantial changes in gene expression, which may be accompanied by post-transcriptional modifications including RNA cytosine-5 methylation (m(5)C). Despite several reports on the transcriptome profiling of activated neurons, the dynamics of neuronal mRNA m(5)C modification in response to environmental stimuli has not been explored. Here, we provide transcriptome-wide maps of m(5)C modification, together with gene expression profiles, for mouse cortical neurons at 0 h, 2 h, and 6 h upon membrane depolarization. Thousands of differentially expressed genes (DEGs) were identified during the neuronal depolarization process. In stimulated neurons, the majority of early response genes were found to serve as expression regulators of late response genes, which are involved in signaling pathways and diverse synaptic functions. With RNA bisulfite sequencing data, a union set of 439 m(5)C sites was identified with high confidence, and approximately 30% of them were shared by neurons at all three time points. Interestingly, over 41% of the m(5)C sites showed increased methylation upon neuronal activation and were enriched in transcripts coding for proteins with synaptic functions. In addition, a modest negative correlation was observed between RNA expression and methylation. In summary, our study provided dynamic transcriptome-wide landscapes of RNA m(5)C methylation in neurons, and revealed that mRNA m(5)C methylation is associated with the regulation of gene expression.