N6-methyladenosine modification of mRNAs in retinal ischemia-reperfusion in mice

Retinal ischemia-reperfusion injury (RIR) is the main pathogenic mechanisms of acute glaucoma, diabetic retinopathy, central retinal vein occlusion. As a common post-transcriptional modification of eukaryotic RNAs, N6-methyladenosine (m⁶A) is associated with the pathogenesis of different diseases, including angiogenesis, through the regulation of RNA metabolism and functions. The aim of this study was to identify the potential relevance of m⁶A RNA methylation in pathogenesis of RIR. A total of 10,851 mRNAs and 23,270 associated m⁶A methylation modified peaks were identified in the RIR group. Similarly, 10,391 mRNAs and 22,935 associated m⁶A methylation modified peaks were detected in the Sham group. MeRIP-seq identified 3,871 RIR-specific m⁶A peaks and 3,624 Sham-specific m⁶A peaks, in addition to 19,399 shared peaks between groups. Gene ontology (GO) analysis showed that hypermethylated mRNAs were enriched in cellular process, cellular anatomical entity, and binding, while hypomethylated mRNAs were enriched in synaptic signaling, synapse, and gated channel activity. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis indicated that hypermethylated mRNAs were involved in tight junction, hippo signaling pathway, and PI3K-Akt signaling pathway, while hypomethylated mRNAs were involved in Neuroactive ligand-receptor interaction, glutamatergic synapses, cholinergic synapses. Joint analysis identified mRNAs with differential m⁶A methylation and expression simultaneously. Among them, the expression patterns of Irx4, Kdr, and Lyz2 were confirmed by RT-qPCR to be consistent with the sequencing results. The results revealed an altered m⁶A epitranscriptome in RIR retinas. These methylated RNAs may act as novel modulators and targets in RIR.