The expressions and m⁶A methylation modification patterns of mRNAs and lncRNAs in major depressive disorder

To systematically characterize the epitranscriptomic landscape of N⁶-methyladenosine (m⁶A) methylation in major depressive disorder (MDD) and elucidate its functional consequences on mRNA and lncRNA regulation.

We performed comprehensive m⁶A methylome profiling using microarray analysis of peripheral blood samples from first-episode, treatment-naïve MDD patients (n = 4) and matched healthy controls (n = 4). Differentially methylated RNAs were identified through stringent criteria (fold-change ≥1.5, probability value (p) < 0.05), followed by integrated bioinformatics analyses including Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis. Validation of selected RNAs (GRM4, CAMKK2, TTC28-AS1, and PAX8-AS1) was performed using MeRIP-qPCR in 15 MDD patients and 15 controls.

Significant differences in m⁶A methylation of mRNAs and lncRNAs were observed between MDD patients and healthy controls. 513 mRNAs (180 hypermethylated, 333 hypomethylated) and 119 lncRNAs (58 hypermethylated, 61 hypomethylated) showed differential m⁶A modifications. GO and KEGG analyses revealed that hypermethylated mRNAs were mainly involved in neuroactive ligand-receptor interactions, while hypomethylated mRNAs were associated with the AMPK signaling pathway. Among the 350 differentially expressed mRNAs, 171 were upregulated and 179 downregulated, enriched in cAMP and TNF signaling pathways, respectively. Among 110 differentially expressed lncRNAs, 71 were upregulated and 39 downregulated. Validation confirmed significant m⁶A methylation differences in GRM4 (p < 0.001), CAMKK2 (p = 0.012), and PAX8-AS1 (p = 0.006), but not significantly different in TTC28-AS1.

Our findings suggest that peripheral m⁶A methylation signatures are associated with MDD pathophysiology, with GRM4 and PAX8-AS1 representing promising candidate biomarkers, while the AMPK and neuroactive ligand-receptor pathways provide hypothesis-generating avenues for future mechanistic investigation.