Identification of the rabies virus alpha/beta interferon antagonist: phosphoprotein P interferes with phosphorylation of interferon regulatory factor 3.
Rabies virus (RV) of the Rhabdoviridae family grows in alpha/beta interferon (IFN)-competent cells, suggesting the existence of viral mechanisms preventing IFN gene expression. We here identify the viral phosphoprotein P as the responsible IFN antagonist. The critical involvement of P was first suggested by the observation that an RV expressing an enhanced green fluorescent protein (eGFP)-P fusion protein (SAD eGFP-P) (S. Finke, K. Brzozka, and K. K. Conzelmann, J. Virol. 78:12333-12343, 2004) was eliminated in IFN-competent HEp-2 cell cultures, in contrast to wild-type (wt) RV or an RV replicon lacking the genes for matrix protein and glycoprotein. SAD eGFP-P induced transcription of the IFN-beta gene and expression of the IFN-responsive MxA and STAT-1 genes. Similarly, an RV expressing low levels of P, which was generated by moving the P gene to a promoter-distal gene position (SAD DeltaPLP), lost the ability to prevent IFN induction. The analysis of RV mutants lacking expression of truncated P proteins P2, P3, or P4, which are expressed from internal AUG codons of the wt RV P open reading frame, further showed that full-length P is competent in suppressing IFN-beta gene expression. In contrast to wt RV, the IFN-inducing SAD DeltaPLP caused S386 phosphorylation, dimerization, and transcriptional activity of IFN regulatory factor 3 (IRF-3). Phosphorylation of IRF-3 by TANK-binding kinase-1 expressed from transfected plasmids was abolished in wt RV-infected cells or by cotransfection of P-encoding plasmids. Thus, RV P is necessary and sufficient to prevent a critical IFN response in virus-infected cells by targeting activation of IRF-3 by an upstream kinase.
Brzózka
Krzysztof
K
Max-von-Pettenkofer Institute & Gene Center, Feodor-Lynen-Str. 25, D-81377 Munich, Germany.
Finke
Stefan
S
Conzelmann
Karl-Klaus
KK
D000818:Animals; D002460:Cell Line; D002522:Chlorocebus aethiops; D004268:DNA-Binding Proteins / Q000378:metabolism*; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D050838:Interferon Regulatory Factor-3; D016898:Interferon-alpha / Q000037:antagonists & inhibitors* / Q000378:metabolism; D016899:Interferon-beta / Q000037:antagonists & inhibitors* / Q000378:metabolism; D018832:Molecular Chaperones; D010750:Phosphoproteins / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D011820:Rabies virus / Q000235:genetics / Q000378:metabolism / Q000472:pathogenicity*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D014157:Transcription Factors / Q000378:metabolism*; D015533:Transcriptional Activation / Q000187:drug effects*; D014709:Vero Cells; D015678:Viral Structural Proteins / Q000235:genetics / Q000378:metabolism*
D004268:DNA-Binding Proteins; C494232:IRF3 protein, human; D050838:Interferon Regulatory Factor-3; D016898:Interferon-alpha; D018832:Molecular Chaperones; C103961:P phosphoprotein, Rabies virus; D010750:Phosphoproteins; D011993:Recombinant Fusion Proteins; D014157:Transcription Factors; D015678:Viral Structural Proteins; D049452:Green Fluorescent Proteins; D016899:Interferon-beta; D017346:Protein Serine-Threonine Kinases; C403191:TBK1 protein, human