Virulent Poxviruses Inhibit DNA Sensing by Preventing STING Activation.

Iliana Georgana,Rebecca P Sumner,Greg J Towers,Carlos Maluquer De Motes,Jae U Jung

doi:10.1128/jvi.02145-17

Iliana Georgana, Rebecca P Sumner + Show 3 more

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https://doi.org/10.1128/jvi.02145-17

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Abstract

ABSTRACTCytosolic recognition of DNA has emerged as a critical cellular mechanism of host immune activation upon pathogen invasion. The central cytosolic DNA sensor cGAS activates STING, which is phosphorylated, dimerizes and translocates from the endoplasmic reticulum (ER) to a perinuclear region to mediate IRF-3 activation. Poxviruses are double-stranded DNA viruses replicating in the cytosol and hence likely to trigger cytosolic DNA sensing. Here, we investigated the activation of innate immune signaling by 4 different strains of the prototypic poxvirus vaccinia virus (VACV) in a cell line proficient in DNA sensing. Infection with the attenuated VACV strain MVA activated IRF-3 via cGAS and STING, and accordingly STING dimerized and was phosphorylated during MVA infection. Conversely, VACV strains Copenhagen and Western Reserve inhibited STING dimerization and phosphorylation during infection and in response to transfected DNA and cyclic GMP-AMP, thus efficiently suppressing DNA sensing and IRF-3 activation. A VACV deletion mutant lacking protein C16, thought to be the only viral DNA sensing inhibitor acting upstream of STING, retained the ability to block STING activation. Similar inhibition of DNA-induced STING activation was also observed for cowpox and ectromelia viruses. Our data demonstrate that virulent poxviruses possess mechanisms for targeting DNA sensing at the level of the cGAS-STING axis and that these mechanisms do not operate in replication-defective strains such as MVA. These findings shed light on the role of cellular DNA sensing in poxvirus-host interactions and will open new avenues to determine its impact on VACV immunogenicity and virulence.IMPORTANCE Poxviruses are double-stranded DNA viruses infecting a wide range of vertebrates and include the causative agent of smallpox (variola virus) and its vaccine vaccinia virus (VACV). Despite smallpox eradication VACV remains of interest as a therapeutic. Attenuated strains are popular vaccine candidates, whereas replication-competent strains are emerging as efficient oncolytics in virotherapy. The successful therapeutic use of VACV depends on a detailed understanding of its ability to modulate host innate immune responses. DNA sensing is a critical cellular mechanism for pathogen detection and activation of innate immunity that is centrally coordinated by the endoplasmic reticulum-resident protein STING. Here, STING is shown to mediate immune activation in response to MVA, but not in response to virulent VACV strains or other virulent poxviruses, which prevent STING activation and DNA sensing during infection and after DNA transfection. These results provide new insights into poxvirus immune evasion and have implications in the rational design of VACV-based therapeutics.

Highlights

Cytosolic recognition of DNA has emerged as a critical cellular mechanism of host immune activation upon pathogen invasion
To seek evidence for such inhibitors, we investigated the differential capacities of four vaccinia virus (VACV) strains to inhibit innate immunity and stimulator of interferon genes (STING) activation in a human monocytic cell line in response to DNA sensing
Similar to the results obtained in cells depleted for STING, Modified vaccinia virus Ankara (MVA) infection triggered a significantly reduced response in cells depleted for cyclic GMP-AMP synthase (cGAS) (Fig. 2F). These results demonstrate that cGAS and STING contribute to IFN responsive factors (IRFs)-3 activation in THP-1 cells infected with MVA

Summary

Introduction

Cytosolic recognition of DNA has emerged as a critical cellular mechanism of host immune activation upon pathogen invasion. Cytosolic double-stranded RNA is sensed by retinoic acid-inducible gene I (RIG-I) and melanoma differentiationassociated gene 5 (MDA-5) Both RIG-I and MDA-5 associate with the IFN-␤ promoter stimulator 1 (IPS-1; known as MAVS), a protein residing in the mitochondrion that mediates the activation of IFN responsive factors (IRFs) and nuclear factor ␬-light-chainenhancer of activated B cells (NF-␬B) [1]. Besides cGAS, other molecules have been proposed to recognize cytosolic DNA and contribute to STING-dependent IFN responses, including DAI [9]; IFI16 [10]; the DExD/ H-box helicases DHX9, DDX36, [11], and DDX41 [12]; and the DNA damage proteins Ku70/80 [13], DNA-PK [14], and Mre11 [15] Whether and how these molecules impact the cGAS-cGAMP-STING axis and whether they show pathogen and/or cell type specificity are important questions in the field [16, 17]. In contrast to COP or WR, which retain full inhibitory capacity, MVA has been shown to trigger innate immune activation in multiple experimental settings

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