title: 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.
authors.affiliation: Laboratory of Molecular Virology and Biotechnology, Institute of Biotechnology, National Cheng Kung University, Tainan 701, Taiwan, ROC.
authors.forename: Yu-Chin
authors.initials: YC
authors.lastname: Su
authors.forename: Jen-Leih
authors.initials: JL
authors.lastname: Wu
authors.forename: Jiann-Ruey
authors.initials: JR
authors.lastname: Hong
mesh_terms: D051702:Activating Transcription Factor 6 / Q000378:metabolism; D000818:Animals; D016923:Cell Death; D002460:Cell Line; D002470:Cell Survival; D004268:DNA-Binding Proteins / Q000378:metabolism; D012321:DNA-Directed RNA Polymerases / Q000378:metabolism; D004721:Endoplasmic Reticulum / Q000378:metabolism / Q000473:pathology*; D000091342:Endoplasmic Reticulum Chaperone BiP; D005399:Fishes / Q000821:virology*; D055785:Gene Knockdown Techniques; D006360:Heat-Shock Proteins / Q000378:metabolism*; D008928:Mitochondria / Q000378:metabolism*; D030381:Nodaviridae / Q000502:physiology*; D010766:Phosphorylation; D011485:Protein Binding; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D012367:RNA, Viral / Q000378:metabolism; D000071701:Regulatory Factor X Transcription Factors; D013312:Stress, Physiological; D014157:Transcription Factors / Q000378:metabolism; D015854:Up-Regulation; D019562:Viral Load; D014764:Viral Proteins / Q000378:metabolism; D014779:Virus Replication / Q000502:physiology*; D019892:eIF-2 Kinase / Q000378:metabolism
chemical_list: D051702:Activating Transcription Factor 6; D004268:DNA-Binding Proteins; D012321:DNA-Directed RNA Polymerases; D000091342:Endoplasmic Reticulum Chaperone BiP; D006360:Heat-Shock Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D012367:RNA, Viral; D000071701:Regulatory Factor X Transcription Factors; D014157:Transcription Factors; D014764:Viral Proteins; D019892:eIF-2 Kinase