Deoxyguanosine Kinase Deficiency Couples Purine Metabolism to Innate Immune Activation and Lipid Accumulation in Hepatocytes

Mitochondrial DNA depletion syndromes (MDS) encompass a heterogeneous set of metabolic disorders caused by defects in enzymes responsible for maintaining mitochondrial nucleotide pools and genome integrity. Among these, deoxyguanosine kinase (DGUOK) acts within the mitochondrial purine-salvage pathway and loss-of-function mutations give rise to DGUOK deficiency, a severe hepatocerebral form of MDS marked by liver failure, neurodevelopmental impairment, and systemic metabolic inflammation. Although the clinical manifestations of DGUOK deficiency have been primarily ascribed to defective mitochondrial DNA (mtDNA) replication, some patients exhibit hepatic steatosis and inflammation despite preserved mtDNA content, suggesting that DGUOK deficiency may deregulate additional metabolic and immune pathways. Here we show that DGUOK depletion reprograms hepatocellular metabolism and innate immune signaling through a purine-dependent mechanism operating independently of mtDNA depletion. In Human hepatocellular carcinoma (HEPG2) hepatocytes subjected to siRNA-mediated DGUOK silencing, mitochondrial architecture and respiration remained intact but cells exhibited pronounced lipid droplet accumulation and a robust cell-intrinsic innate immune type I interferon response. Bulk RNA sequencing revealed widespread transcriptional reprogramming, including upregulation of human endogenous retroviruses (HERVs) and interferon-stimulated genes (ISGs), suppression of lipid metabolic pathways, and changes in purine, methionine, and methylation-associated gene networks. Perturbing purine homeostasis through deoxyadenosine (dAdo) supplementation in wild-type cells phenocopied DGUOK disruption, causing cellular DNA hypomethylation and activation of viral mimicry pathways. Together, these findings identify DGUOK as a central regulator of the purine-regulated lipid-immune axis in hepatocytes, demonstrating that mitochondrial nucleotide salvage preserves hepatic immune and metabolic homeostasis beyond its canonical role in mtDNA synthesis. By linking purine imbalances to steatosis and type I interferon activation, this study establishes a mechanistic framework for immunometabolic pathology in DGUOK deficiency.