Pseudorabies virus is a typical swine alphaherpesvirus, which can cause obvious neurological disorders and reproductive failure in pigs. It is capable of evading host antiviral immune response. However, the mechanism by which many PRV proteins assist the virus to evade innate immunity is not fully understood. This study identified PRV US3 protein as a crucial antagonistic viral factor that represses interferon beta (IFN-β) expression. A in-depth study showed that US3 protein restricted type I IFN production by targeting interferon regulatory factor 3 (IRF3), a key molecule required for type I IFN induction. Additionally, US3 protein interacted with IRF3, degraded its protein expression to block the phosphorylation of IRF3. These findings suggested a novel strategy utilized by PRV to inhibit IFN-β production and escape the host innate immunity.

Interferon (IFN) helps cells fight viral infections by further inducing the expression of many downstream IFN-stimulated genes (ISGs). Human interferon-inducible transmembrane proteins (IFITM) are one of these ISGs. The antiviral function of human IFITM1, IFITM2, and IFITM3 are well known. In this study, we report that IFITM can significantly inhibit EMCV infectivity in HEK293 cells. Overexpression of IFITM proteins could promote IFN-β production. Meanwhile, IFITMs facilitated type I IFN signaling pathway adaptor MDA5 expression. We detected the binding of IFITM2 to MDA5 in a co-immunoprecipitation assay. It was also found that the ability of IFITM2 to activate IFN-β was significantly inhibited after interfering with MDA5 expression, suggesting that MDA5 may play an important role in the activation of the IFN-β signaling pathway by IFITM2. Moreover, the N-terminal domain plays an active role in the antiviral activity and the activation of IFN-β by IFITM2. These findings suggest that IFITM2 plays a vital role in antiviral signaling transduction. In addition, a positive feed-forward loop between IFITM2 and type I IFN establishes a key role for IFITM2 in enforcing innate immune responses.

Pseudorabies (PR) is a domestic and wild animal infectious disease caused by the pseudorabies virus (PRV) and is one of the major infectious diseases that endanger the global swine industry. Studies have reported that PRV may achieve cross-species transmission from pigs to humans in recent years. Therefore, in-depth exploration of the relationship between PRV and host proteins is of great significance for elucidating the pathogenic mechanism of PRV and anti-PRV infection. Here, we report that heat shock protein 27 (HSP27) ubiquitinates and degrades cyclic GMP-AMP synthase (cGAS) and attenuates cGAS-mediated antiviral responses, thereby promoting PRV infection. Overexpression of HSP27 promoted PRV proliferation in vitro, while knockdown of HSP27 inhibited PRV infection. Importantly, we found that HSP27 inhibited PRV infection or poly(dA:dT)-activated IFN-β expression. Further studies found that HSP27 may inhibit cGAS-STING-mediated IFN-β expression through targeting cGAS. In addition, we found that HSP27 can suppress the expression of endogenous cGAS in different cells at both gene transcription and protein expression levels, and that HSP27 interacts with and ubiquitinates cGAS. In conclusion, we reveal for the first time that HSP27 is a novel negative regulator of the cGAS-STING signaling pathway induced by PRV infection or poly(dA:dT) activation and demonstrate that HSP27 plays a crucial role in PRV infection.

Herpesviruses antagonize host antiviral responses through a myriad of molecular strategies culminating in the death of the host cells. Pseudorabies virus (PRV) is a significant veterinary pathogen in pigs, causing neurological sequalae that ultimately lead to the animal's demise. PRV is known to trigger apoptotic cell death during the late stages of infection. The virion host shutdown protein (VHS) encoded by UL41 plays a crucial role in the PRV infection process. In this study, we demonstrate that UL41 inhibits PRV-induced activation of inflammatory cytokine and negatively regulates the cGAS-STING-mediated antiviral activity by targeting IRF3, thereby inhibiting the translocation and phosphorylation of IRF3. Notably, mutating the conserved amino acid sites (E192, D194, and D195) in the RNase domain of UL41 or knocking down UL41 inhibits the immune evasion of PRV, suggesting that UL41 may play a crucial role in PRV's evasion of the host immune response during infection. These results enhance our understanding of how PRV structural proteins assist the virus in evading the host immune response.

Nuclear factor κB (NF-κB) is involved in a wide range of innate immune activities in host cells and serves as an important component of a host’s immunity system. To survive in infected cells, viruses have evolved intricate strategies to evade the host immune response. Pseudorabies virus (PRV) is a member of the alpha herpesvirus family and is capable of causing reproductive and neurological dysfunction in pigs. PRV has a large DNA genome and therefore has the ability to encode numerous proteins that modulate host innate immune responses. In the present study, we demonstrated that the PRV-encoded immediate early protein ICP0 inhibits the tumor necrosis factor alpha (TNF-α)-mediated NF-κB signaling pathway. An in-depth study showed that ICP0 protein was able to limit NF-κB activation and decreased the expression of inflammatory cytokines interleukin-6 (IL-6) and interleukin 8 (IL-8). In addition, ICP0 blocked the activation of NF-κB through interacting with p65, degrading its protein expression and limiting its phosphorylation. PRV protein ICP0 is shown for the first time to enable escape from innate immune response through the regulation of NF-κB during PRV infection. These results illustrate that PRV ICP0 is able to block NF-κB activation. This mechanism may represent a critical role in the early events leading to PRV infection.

Interferon (IFN)-γ inducible protein 16 (IFI16), a key DNA sensor, triggers downstream STING-dependent type I interferon (IFN-I) production and antiviral immunity. However, how the IFI16-STING signaling pathway is regulated by EMCV infection is still not well elucidated. In this study, we investigated the interaction between IFI16 and EMCV. Results indicated EMCV infection suppressed IFI16 expression in A549 cells. This study reveals that IFI16 plays an active role in combating EMCV. Screening viral proteins in conjunction with IFI16, we found that the EMCV VP2 protein hinders the antiviral response mediated by IFI16 by causing degradation of the IFI16 protein via the caspase-dependent apoptosis pathway. Our study communicates the antiviral role of the IFI16-STING pathway during EMCV infection. Importantly, this study unveils the novel mechanism by which VP2 counteracts the innate immune signaling activated by foreign DNA.