Blocking double-stranded RNA-activated protein kinase PKR by Japanese encephalitis virus nonstructural protein 2A.
Japanese encephalitis virus (JEV) is an enveloped flavivirus with a single-stranded, positive-sense RNA genome encoding three structural and seven nonstructural proteins. To date, the role of JEV nonstructural protein 2A (NS2A) in the viral life cycle is largely unknown. The interferon (IFN)-induced double-stranded RNA (dsRNA)-activated protein kinase (PKR) phosphorylates the eukaryotic translation initiation factor 2α subunit (eIF2α) after sensing viral RNA and results in global translation arrest as an important host antiviral defense response. In this study, we found that JEV NS2A could antagonize PKR-mediated growth inhibition in a galactose-inducible PKR-expressing yeast system. In human cells, PKR activation, eIF2α phosphorylation, and the subsequent translational inhibition and cell death triggered by dsRNA and IFN-α were also repressed by JEV NS2A. Moreover, among the four eIF2α kinases, NS2A specifically blocked the eIF2α phosphorylation mediated by PKR and attenuated the PKR-promoted cell death induced by the chemotherapeutic drug doxorubicin. A single point mutation of NS2A residue 33 from Thr to Ile (T33I) abolished the anti-PKR potential of JEV NS2A. The recombinant JEV mutant carrying the NS2A-T33I mutation showed reduced in vitro growth and in vivo virulence phenotypes. Thus, JEV NS2A has a novel function in blocking the host antiviral response of PKR during JEV infection.
Graduate Institute of Life Sciences, National Defense Medical Center, Taipei, Taiwan.
Yu-Chun
YC
Tu
Chia-Yi
CY
Yu
Jian-Jong
JJ
Liang
Elong
E
Lin
Ching-Len
CL
Liao
Yi-Ling
YL
Lin
D000818:Animals; D002460:Cell Line; D045744:Cell Line, Tumor; D002470:Cell Survival; D004664:Encephalitis Virus, Japanese / Q000378:metabolism*; D004789:Enzyme Activation; D015967:Gene Expression Regulation, Viral; D017930:Genes, Reporter; D057809:HEK293 Cells; D006801:Humans; D051379:Mice; D008810:Mice, Inbred C57BL; D009154:Mutation; D010641:Phenotype; D010957:Plasmids / Q000378:metabolism; D014176:Protein Biosynthesis; D012330:RNA, Double-Stranded / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000378:metabolism; D014764:Viral Proteins / Q000737:chemistry* / Q000378:metabolism; D019892:eIF-2 Kinase / Q000037:antagonists & inhibitors* / Q000378:metabolism
D012330:RNA, Double-Stranded; D014764:Viral Proteins; C057421:virus protein 2A; D019892:eIF-2 Kinase