Betanodavirus up-regulates chaperone GRP78 via ER stress: roles of GRP78 in viral replication and host mitochondria-mediated cell death

Abstract

Whether viral pathogens that induce ER stress responses benefit the host or the virus remains controversial. In this study we show that betanodavirus induced ER stress responses up-regulate GRP78, which regulates the viral replication and host cellular mitochondrial-mediated cell death. Betanodavirus (redspotted grouper nervous necrosis virus, RGNNV) infection resulted in the following increased ER stress responses in fish GF-1 grouper fin cells: (1) IRE-1 and ATF-6 sensors at 48 h post-infection (p.i.) that up-regulated chaperone protein GRP78; (2) activation of caspase-12; and (3) PERK phosphorylation and down-regulation of Bcl-2. Analyses of GRP78 functions during viral replication using either loss-of-function or gain-of-function approaches showed that GRP78 over-expression also enhanced viral replication and induced cell death. Then, we found that zfGRP78 localization gradually increased in mitochondria after RGNNV infection by EGFP tagging approach. Furthermore, zfGRP78 can interact with viral RNA-dependent RNA polymerase (RdRp) by using immunofluorescent and immunoprecipitation assays. Finally, we found that blocking GRP78-mediated ER signals can reduce the viral death factors protein α and protein B2 expression and decrease the Bcl-2 down-regulation mediated mitochondria-dependent cell death, which also enhances host cellular viability. Taken together, our results suggest that RGNNV infection and expression can trigger ER stress responses, which up-regulate the chaperone GRP78 at early replication stage. Then, GRP78 can interact with RdRp that may enhance the viral replication for increasing viral death factors' expressions at middle-late replication stage, which can enhance mitochondrial-mediated cell death pathway and viral spreading. These results may provide new insights into the mechanism of ER stress-mediated cell death in RNA viruses.

Ameliorating ER-stress attenuates Aeromonas hydrophila-induced mitochondrial dysfunctioning and caspase mediated HKM apoptosis in Clarias batrachus

“…Since, A. hydrophila displays wide host specificity it would be interesting to see whether same pathogenic mechanisms are employed across other host species. Microbe-induced ER-stress and UPR has been reported in fish37 and mammals3638394041. Our study not only extends these findings to A. hydrophila but further suggests UPR to be an evolutionarily conserved pathological mechanism.…”

“…While eIF2α halts translation of new proteins in ER and is implicated in apoptosis, BiP acts as pro-survival factor under different stress conditions36. Earlier reports implicated ER-stress and BiP expression to be critical in betanodavirus-induced apoptosis in fish cells37. We wondered the elevated expression of the three ER-stress markers have any correlation with A. hydrophila -induced HKM apoptosis.…”

LipoxinA4 attenuates myocardial ischemia reperfusion injury via a mechanism related to downregulation of GRP-78 and caspase-12 in rats

“…UPR is an evolutionarily conserved response that can be activated to enhance the protein-folding capacity of the ER and promote ER-associated protein degradation to remove the misfolded proteins. Therefore, the release of GRP-78 is critical in the early protective response, which is an upstream gene in the ERS signaling and can also be seen as a marker in the early ERS [15]. In our study, the increased mRNA and protein expression of GRP-78 induced by I/R demonstrated the process of ERS, which can protect the ER and further maintain ER homeostasis.…”

Betanodavirus: Mitochondrial disruption and necrotic cell death

“…In contrast, the betanodavirus B2 protein appears to function as a suppressor of host siRNA silencing [12,13] or as a EDITORIAL necrotic death factor [14,15] . In addition, red-spotted grouper nervous necrosis virus (RGNNV) infection and expression can trigger the ER stress response, which results in the upregulation of the 78 kDa glucose regulated protein at the early replication stage [16] . Very recently, RGNNV has been shown to induce the production of reactive oxygen species (ROS) during the early and middle replication stages [17] .…”

“…In our fish system, we found that RGNNV infection can induce downregulation of the anti-apoptotic Bcl-2 genes at the middle apoptotic stage (48 h p.i.) [16] . Subsequently, mitochondrial damage and RGNNV-induced necrotic cell death were assessed in stable cell lines producing the anti-apoptotic Bcl-2 proteins, zfBcl-xL or zfMcl-1a.…”