A Luciferase Reporter Gene Assay to Measure Ebola Virus Viral Protein 35-Associated Inhibition of Double-Stranded RNA-Stimulated, Retinoic Acid-Inducible Gene 1-Mediated Induction of Interferon β.
During Ebola virus (EBOV) infection, the type I interferon α/β (IFN-α/β) innate immune response is suppressed by EBOV viral protein 35 (VP35), a validated drug target. Identification of EBOV VP35 inhibitors requires a cellular system able to assess the VP35-based inhibitory functions of viral double-stranded RNA (dsRNA) IFN-β induction. We established a miniaturized luciferase gene reporter assay in A549 cells that measures IFN-β induction by viral dsRNA and is dose-dependently inhibited by VP35 expression. When compared to influenza A virus NS1 protein, EBOV VP35 showed improved inhibition of viral dsRNA-based IFN-β induction. This assay can be used to screen for EBOV VP35 inhibitors.
Department of Life and Environmental Sciences, University of Cagliari, Italy.
Valeria
V
Cannas
Department of Life and Environmental Sciences, University of Cagliari, Italy.
Gian Luca
GL
Daino
Department of Life and Environmental Sciences, University of Cagliari, Italy.
Angela
A
Corona
Department of Life and Environmental Sciences, University of Cagliari, Italy.
Francesca
F
Esposito
Department of Life and Environmental Sciences, University of Cagliari, Italy.
Enzo
E
Tramontano
D002460:Cell Line; D029043:Ebolavirus / Q000235:genetics*; D017930:Genes, Reporter / Q000235:genetics*; D019142:Hemorrhagic Fever, Ebola / Q000235:genetics* / Q000821:virology; D006801:Humans; D016898:Interferon-alpha / Q000235:genetics; D016899:Interferon-beta / Q000235:genetics*; D008156:Luciferases / Q000378:metabolism*; D012330:RNA, Double-Stranded / Q000235:genetics*; D015533:Transcriptional Activation / Q000235:genetics; D014212:Tretinoin / Q000378:metabolism; D017361:Viral Nonstructural Proteins / Q000235:genetics; D014764:Viral Proteins / Q000235:genetics / Q000378:metabolism*
C085820:INS1 protein, influenza virus; D016898:Interferon-alpha; D012330:RNA, Double-Stranded; D017361:Viral Nonstructural Proteins; D014764:Viral Proteins; D014212:Tretinoin; D016899:Interferon-beta; D008156:Luciferases
Ebola virus; IFN-β induction; IFN-β inhibition; NS1; RIG-I activation; VP35; drug development; dsRNA recognition; innate immunity; luciferase reporter gene assay