Research ArticleImmunology

Pinpointing cysteine oxidation sites by high-resolution proteomics reveals a mechanism of redox-dependent inhibition of human STING

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Science Signaling  27 Apr 2021:
Vol. 14, Issue 680, eaaw4673
DOI: 10.1126/scisignal.aaw4673

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Taking out the STING

The adaptor protein STING is stimulated in response to the detection of cytosolic viral DNA by the enzyme cGAS. STING activation then leads to the expression of genes encoding type I interferons as part of the antiviral response. STING activation also plays a role in antitumor immunity, making it an important therapeutic target. Zamorano Cuervo et al. used mass spectrometry and structural analyses to identify cysteine residues in STING that underwent reversible oxidation. One specific residue was oxidized in response to oxidants or a natural STING agonist, which the authors propose limits STING activity. Together, these findings may help in the design of therapeutics to modulate STING activity in the context of immunotherapies and vaccines.

Abstract

Protein function is regulated by posttranslational modifications (PTMs), among which reversible oxidation of cysteine residues has emerged as a key regulatory mechanism of cellular responses. Given the redox regulation of virus-host interactions, the identification of oxidized cysteine sites in cells is essential to understand the underlying mechanisms involved. Here, we present a proteome-wide identification of reversibly oxidized cysteine sites in oxidant-treated cells using a maleimide-based bioswitch method coupled to mass spectrometry analysis. We identified 2720 unique oxidized cysteine sites within 1473 proteins with distinct abundances, locations, and functions. Oxidized cysteine sites were found in numerous signaling pathways, many relevant to virus-host interactions. We focused on the oxidation of STING, the central adaptor of the innate immune type I interferon pathway, which is stimulated in response to the detection of cytosolic DNA by cGAS. We demonstrated the reversible oxidation of Cys148 and Cys206 of STING in cells. Molecular analyses led us to establish a model in which Cys148 oxidation is constitutive, whereas Cys206 oxidation is inducible by oxidative stress or by the natural ligand of STING, 2′3′-cGAMP. Our data suggest that the oxidation of Cys206 prevented hyperactivation of STING by causing a conformational change associated with the formation of inactive polymers containing intermolecular disulfide bonds. This finding should aid the design of therapies targeting STING that are relevant to autoinflammatory disorders, immunotherapies, and vaccines.

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