Macrophage RSAD2 Couples mtDNA Synthesis With a Self‐Amplifying Inflammatory Circuit to Orchestrate Tissue Repair

It is well established that mitochondrial DNA (mtDNA) synthesis governs macrophage function, yet its role in tissue repair and inflammation remains poorly understood. Using a mouse skin injury model combined with spatial transcriptomics and functional assays, we identify radical s-adenosyl methionine domain containing 2 (RSAD2) as a critical regulator of mtDNA-driven inflammation in macrophages. Mechanistically, RSAD2 directly binds the N-terminal domain of cytidylate monophosphate kinase 2 (CMPK2), a rate-limiting enzyme for mtDNA synthesis, and dually modulates its activity: it inhibits K63-linked ubiquitination to stabilize CMPK2, and recruits casein kinase 2 alpha 2 (Csnk2a2) to promote CMPK2 phosphorylation, thereby enhancing mtDNA production. Newly synthesized mtDNA amplifies inflammation through two coordinated pathways: activation of a cGAS-STING-IRF3-RSAD2 feedforward loop, and synergistic activation of the NLR family pyrin domain containing 3 (NLRP3) inflammasome. These pathways orchestrate inflammatory amplification in macrophages to modulate skin repair. Our work establishes macrophage RSAD2 as a central hub that coordinates two mtDNA-dependent inflammatory axes, revealing a feedforward immuno-metabolic circuit with broad implications for inflammation-driven diseases.