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

Abstract

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.