Transient focal ischemia significantly alters the m ⁶A epitranscriptomic tagging of RNAs in the brain

Abstract

Background and Purpose

Adenosine in many types of RNAs can be converted to N ⁶-methyladenosine (m ⁶A) which is a highly dynamic epitranscriptomic modification that regulates RNA metabolism and function. Of all organs, the brain shows the highest abundance of m ⁶A methylation of RNAs. As recent studies showed that m ⁶A modification promotes cell survival after adverse conditions, we currently evaluated the effect of stroke on cerebral m ⁶A methylation in mRNAs and lncRNAs.

Methods

Adult C57BL/6J mice were subjected to transient middle cerebral artery occlusion. In the peri-infarct cortex, m ⁶A levels were measured by dot blot analysis and transcriptome-wide m ⁶A changes were profiled using immunoprecipitated methylated RNAs with microarrays (44,122 mRNAs and 12,496 lncRNAs). Gene ontology analysis was conducted to understand the functional implications of m ⁶A changes after stroke. Expression of m ⁶A writers, readers and erasers was also estimated in the ischemic brain.

Results

Global m ⁶A levels increased significantly at 12h and 24h of reperfusion compared to sham. While 139 transcripts (122 mRNAs and 17 lncRNAs) were hypermethylated, 8 transcripts (5 mRNAs and 3 lncRNAs) were hypomethylated (>5-fold compared to sham) in the ischemic brain at 12h reperfusion. Inflammation, apoptosis and transcriptional regulation are the major biological processes modulated by the post-stroke differentially m ⁶A methylated mRNAs. The m ⁶A writers were unaltered, but the m ⁶A eraser (fat mass and obesity-associated protein) decreased significantly after stroke compared to sham.

Conclusions

This is the first study to show that stroke alters the cerebral m ⁶A epitranscriptome which might have functional implications in post-stroke pathophysiology.