METTL3-Mediated N⁶-methyladenosine mRNA Modification and cGAS-STING Pathway Activity in Kidney Fibrosis

Background Chemical modifications on RNA profoundly impact RNA function and regulation. m(6)A, the most abundant RNA modification in eukaryotes, plays a pivotal role in diverse cellular processes and disease mechanisms. However, its importance is understudied in human chronic kidney disease (CKD) samples regarding its influence on pathological mechanisms. Methods LC-MS/MS and Methylated RNA Immunoprecipitation (MeRIP) sequencing were utilized to examine alterations in m(6)A levels and patterns in CKD samples. Overexpression of the m(6)A writer METTL3 in cultured kidney tubular cells was performed to confirm the impact of m(6)A in tubular cells and explore the biological functions of m(6)A modification on target genes. Additionally, tubule-specific deletion of Mettl3 (Ksp-Cre Mettl3(f/f)) mice and the use of anti-sense oligonucleotides inhibiting Mettl3 expression were utilized to reduce m(6)A modification in an animal kidney disease model. Results By examining 127 human CKD samples, we observed a significant increase in m(6)A modification and METTL3 expression in diseased kidneys. Epitranscriptomic analysis unveiled an enrichment of m(6)A modifications in transcripts associated with the activation of inflammatory signaling pathways, particularly the cGAS-STING pathway. m(6)A hypermethylation increased mRNA stability in cGAS and STING1, as well as elevated the expression of key proteins within the cGAS-STING pathway. Both the tubule-specific deletion of Mettl3 and the use of anti-sense oligonucleotides to inhibit Mettl3 expression protected mice from inflammation, reduced cytokine expression, decreased immune cell recruitment, and attenuated kidney fibrosis. Conclusions Our research revealed heightened METTL3-mediated m(6)A modification in fibrotic kidneys, particularly enriching the cGAS-STING pathway. This hypermethylation increased mRNA stability for cGAS and STING1, leading to sterile inflammation and fibrosis.