Mitochondria serve as the cellular "power plants," supplying energy and regulating metabolism, signal transduction, and other physiological processes. To successfully replicate within host cells, viruses have evolved multiple strategies to hijack mitochondrial functions. The oncolytic Newcastle disease virus (NDV) causes severe organelle damage in tumor cells; however, how it manipulates mitochondrial architecture to facilitate its own replication remains poorly understood. Here, we provide evidence that NDV infection disrupts mitochondrial spatial distribution and imbalances mitochondrial fusion and fission, leading to mitochondrial structural damage. The resulting accumulation of fragmented mitochondria is cleared via PRKN-dependent mitophagy, a process that supports NDV replication. Interestingly, although MAVS (mitochondrial antiviral signaling protein) is degraded along with mitophagy, genetic ablation of PRKN - while blocking MAVS degradation - does not restore downstream innate immune responses. This indicates that NDV exploits mitophagy to enhance replication through mechanisms not entirely dependent on the suppression of MAVS-mediated immunity. Given the central role of mitochondria, we further explored the link between amino acid metabolism and viral proliferation after NDV infection. Our results show that NDV-induced mitophagy leads to the accumulation of free amino acids in host cells, and this metabolic reprogramming promotes viral replication. In summary, we show that NDV drives its replication by remodeling mitochondrial dynamics to induce mitophagy, which in turn triggers an amino acid metabolic reprogramming that benefits the virus. This provides new insights into the mechanisms supporting efficient oncolytic NDV replication, offering potential avenues for therapeutic intervention in oncolytic virus therapy.CCCP: carbonyl cyanide m-chlorophenylhydrazone; COX4/COX IV: cytochrome c oxidase subunit 4; CQ: chloroquine; DENV: dengue virus; DNM1L/DRP1: dynamin 1 lik;ETC: electron transport chain; FIS1: fission, mitochondrial 1; HBV: hepatitis B virus; IAV: influenza A virus; IMM: inner mitochondrial membrane; JEV: japanese encephalitis virus; MAVS: mitochondrial antiviral signaling protein; MFF: mitochondrial fission factor; MFN1: mitofusin 1; MFN2: mitofusin 2; MOI: multiplicity of infection; MV: measles virus; NDV: Newcastle disease virus; OMM: outer mitochondrial membrane; OPA1: OPA1 mitochondrial dynamin like GTPase; PINK1: PTEN induced kinase 1; PRKN/parkin: parkin RBR E3 ubiquitin protein ligase; RLR: RIG-I-like receptor; SDHA: succinate dehydrogenase complex flavoprotein subunit A; TCA: tricarboxylic acid cycle; TCID: tissue culture infective doses; TEM: transmission electron microscopy; TIMM23: translocase of inner mitochondrial membrane 23; TOMM20: translocase of outer mitochondrial membrane 20. Abbreviations: 50
