Induction of Noxa-Mediated Apoptosis by Modified Vaccinia Virus Ankara Depends on Viral Recognition by Cytosolic Helicases, Leading to IRF-3/IFN-β-Dependent Induction of Pro-Apoptotic Noxa

Pedro Eitz Ferrer,Ingo Drexler,Wolfgang Kastenmüller,Stephanie Potthoff,Georg Häcker,Stefan A Paschen,Georg Gasteiger,Holger Ludwig,Gerd Sutter,Andreas Villunger,Susanne Kirschnek

doi:10.1371/journal.ppat.1002083

Abstract

Viral infection is a stimulus for apoptosis, and in order to sustain viral replication many viruses are known to carry genes encoding apoptosis inhibitors. F1L, encoded by the orthopoxvirus modified vaccinia virus Ankara (MVA) has a Bcl-2-like structure. An MVA mutant lacking F1L (MVAΔF1L) induces apoptosis, indicating that MVA infection activates and F1L functions to inhibit the apoptotic pathway. In this study we investigated the events leading to apoptosis upon infection by MVAΔF1L. Apoptosis largely proceeded through the pro-apoptotic Bcl-2 family protein Bak with some contribution from Bax. Of the family of pro-apoptotic BH3-only proteins, only the loss of Noxa provided substantial protection, while the loss of Bim had a minor effect. In mice, MVA preferentially infected macrophages and DCs in vivo. In both cell types wt MVA induced apoptosis albeit more weakly than MVAΔF1L. The loss of Noxa had a significant protective effect in macrophages, DC and primary lymphocytes, and the combined loss of Bim and Noxa provided strong protection. Noxa protein was induced during infection, and the induction of Noxa protein and apoptosis induction required transcription factor IRF3 and type I interferon signalling. We further observed that helicases RIG-I and MDA5 and their signalling adapter MAVS contribute to Noxa induction and apoptosis in response to MVA infection. RNA isolated from MVA-infected cells induced Noxa expression and apoptosis when transfected in the absence of viral infection. We thus here describe a pathway leading from the detection of viral RNA during MVA infection by the cytosolic helicase-pathway, to the up-regulation of Noxa and apoptosis via IRF3 and type I IFN signalling.

Highlights

Cell death by apoptosis can protect a multicellular organism against viral infection: if the first infected cell dies in time the virus will not have the chance of producing new viral particles
By using a broad array of immortalized and primary cell types we observed that viral infection induced programmed cell death was controlled by proteins predominantly involved in detection of viral RNA, in particular proteins involved in the type 1 interferon response
The novelty of our findings lies on the observation that can RNA from DNA viruses be detected and activate the type 1 interferon response to infection, but that these responses can directly modulate the levels of proteins regulating programmed cell death

Summary

Introduction

Cell death by apoptosis can protect a multicellular organism against viral infection: if the first infected cell dies in time the virus will not have the chance of producing new viral particles. Many viruses counter this cellular response by carrying genes coding for inhibitors of the host cell’s apoptosis machinery. A number of poxviruses carry, among other anti-apoptotic genes, genes coding for proteins that resemble the inhibitory members of the mammalian Bcl-2-family of proteins These genes have very little (if any) primary sequence homology with the mammalian proteins but crystallization studies have shown that they adopt a virtually identical structure. One critical feature of anti-apoptotic function of Bcl-2-proteins is the existence of a hydrophobic groove that accommodates the BH3-domain of proapoptotic Bcl-2-family members and thereby inhibits apoptosis [4]

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