Pseudorabies virus pUL40 drives inflammatory signaling through competitive hijacking of EphA2 from the Akt-EphA2 interaction.

Herpesvirus infection triggers excessive inflammation, contributing to tissue injury and disease severity, but the underlying drivers remain unclear. Here we identify EphA2 as a critical mediator of pseudorabies virus (PRV)-induced inflammatory response. Genetic or pharmacological inhibition of EphA2 reduced TNF-α production in PRV-infected cells and mice, alleviated liver and lung damage, and improved mice survival. Mechanistically, PRV infection promoted EphA2 phosphorylation at Ser-897, which activated NF-κB and MAPK pathways to drive TNF-α production. The PRV tegument protein UL40 directly bound the EphA2 kinase domain (residues 697-901), a region that also interacts with Akt. Unexpectedly, Akt acted as a negative regulator of inflammation, as its knockdown exacerbated cytokine production. UL40 competitively disrupted the constitutive EphA2-Akt complex in a time-dependent manner, relieving Akt-mediated restraint on EphA2 S897 phosphorylation. Accordingly, a UL40-deficient PRV mutant failed to enhance S897 phosphorylation, elicited lower inflammatory responses, and showed attenuated virulence in mice. Collectively, our findings reveal a proviral strategy whereby PRV UL40 hijacks EphA2 to counteract an intrinsic Akt-dependent inhibitory pathway. This UL40-EphA2-Akt cascade is a critical determinant of virus-induced inflammation, and EphA2 S897 represents a potential therapeutic target for mitigating alphaherpesvirus immunopathology.