PKM2 Facilitates Classical Swine Fever Virus Replication by Enhancing NS5B Polymerase Function.

Host metabolic reprogramming is a critical strategy employed by many viruses to support their replication, and the key metabolic enzyme plays important roles in virus infection. This study investigates the role of pyruvate kinase M2 (PKM2), a glycolytic enzyme with non-canonical functions, in the replication of classical swine fever virus (CSFV). Using PK-15 cells and piglet models, we demonstrate that CSFV infection upregulates PKM2 expression both in vitro and in vivo, creating a proviral environment. knockdown of PKM2 by siRNA reduced CSFV proliferation, while PKM2 overexpression significantly increased virus propagation, which was evaluated by viral protein synthesis, genome replication, and progeny virion production. A direct interaction between PKM2 and CSFV NS5B protein was identified by co-immunoprecipitation and GST-pulldown assays, and PKM2 affected NS5B polymerase activity in a dual-luciferase reporter assay, with PKM2 depletion reducing RdRp function by 50%. Temporal analysis of the first viral replication cycle confirmed PKM2-dependent enhancement of CSFV RNA synthesis. These findings establish PKM2 as a proviral host factor that directly binds NS5B to potentiate RdRp activity, thereby bridging metabolic adaptation and viral genome replication. This study provides new evidence of a glycolytic enzyme physically interacting and enhancing viral polymerase function, offering new information about CSFV-host interaction.