The potential roles of mA modification in regulating the inflammatory response in microglia.

Background: Microglia are key regulators of the inflammatory response in the brain. Adenosine in RNAs can be converted to mA (N-methyladenosine), which regulates RNA metabolism and functions as a key epitranscriptomic modification. The mA modification pattern and mA-related signatures under pro-inflammatory and anti-inflammatory conditions of microglia remain unclear.

Methods: Primary rat microglia were differentiated into pro-inflammatory M1-like (M1-L), anti-inflammatory M2-like (M2-L), and resting, unstimulated (M0-L) phenotypes. mA mRNA and lncRNA epitranscriptomic microarray analyses were performed, and pathway analysis was conducted to understand the functional implications of mA methylation in mRNAs and lncRNAs. The mA methylation level and gene expression of mRNAs and lncRNAs were subsequently verified by mA Me-RIP and qRT-PCR.

Results: A total of 1588 mRNAs and 340 lncRNAs, 315 mRNAs and 38 lncRNAs, and 521 mRNAs and 244 lncRNAs were differentially mA methylated between M1-L and M0-L (M1-L/M0-L), M2-L and M0-L (M2-L/M0-L), M2-L and M1-L (M2-L/M1-L), respectively. Furthermore, 4902 mRNAs, 4676 mRNAs, and 5095 mRNAs were identified distinctively expressed in M1-L/M0-L, M2-L/M0-L, and M2-L/M1-L, respectively. Pathway analysis of differentially mA methylated mRNAs and lncRNAs in M1-L/M0-L identified immune system, signal transduction, and protein degradation processes. In contrast, the distinct mA methylated mRNAs in M2-L/M0-L were involved in genetic information processing, metabolism, cellular processes, and neurodegenerative disease-related pathways. We validated mA methylation and the expression levels of five mRNAs and five lncRNAs, which were involved in upregulated pathways in M1-L/M0-L, and five mRNAs involved in upregulated pathways in M2-L/M0-L.

Conclusions: These findings identify a distinct mA epitranscriptome in microglia, and which may serve as novel and useful regulator during pro-inflammatory and anti-inflammatory response of microglia.