Porcine reproductive and respiratory syndrome virus Nsp2 induces incomplete autophagy to facilitate viral replication via disrupting SNARE complex assembly

Porcine reproductive and respiratory syndrome (PRRS), which is caused by the PRRS virus (PRRSV), is a highly contagious disease that primarily affects the reproductive and respiratory systems of pigs, severely affecting their health and productivity. Despite extensive research, underlying pathogenic mechanisms remain unclear. In this study, we demonstrated that PRRSV infection triggers incomplete autophagy, which is characterized by impaired autophagosome–lysosome fusion. We screened for viral nonstructural proteins (Nsps) and identified Nsp2 as the key driver of this process. Nsp2 interacts with the SNARE complex proteins STX17 and VAMP8, which are involved in the fusion of autophagosomes and lysosomes. The C-terminal tail domain of Nsp2 (989–1166 aa) binds to the N-terminal region of STX17 (124–166 aa), and the transmembrane domain of VAMP8 interacts with both the PLP2 domain (47–159 aa) and tail domain of Nsp2. Co-immunoprecipitation assays revealed that both PRRSV infection and Nsp2 overexpression competitively inhibited SNARE complex assembly and disrupted SNARE complex formation. To validate the functional link between incomplete autophagy and viral replication, three SNARE-targeting compounds—costunolide (CTD), EACC (ethyl (2-(5-nitrothiophene-2-carboxamido) thiophene-3‑carbonyl), and berbamine (BBM)—were used. CTD and EACC enhanced viral replication efficiency, with CTD exhibiting peak efficacy at 48 h post-infection. These findings demonstrate that PRRSV hijacks autophagy via Nsp2-mediated SNARE disruption to create a replication-favorable niche, providing new insights into antiviral strategies targeting host–pathogen interactions.