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Ubiquitin variants potently inhibit SARS-CoV-2 PLpro and viral replication via a novel site distal to the protease active site.

ABSTRACT: The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic has made it clear that combating coronavirus outbreaks benefits from a combination of vaccines and therapeutics. A promising drug target common to all coronaviruses-including SARS-CoV, MERS-CoV, and SARS-CoV-2-is the papain-like protease (PLpro). PLpro cleaves part of the viral replicase polyproteins into non-structural protein subunits, which are essential to the viral replication cycle. Additionally, PLpro can cleave both ubiquitin and the ubiquitin-like protein ISG15 from host cell substrates as a mechanism to evade innate immune responses during infection. These roles make PLpro an attractive antiviral drug target. Here we demonstrate that ubiquitin variants (UbVs) can be selected from a phage-displayed library and used to specifically and potently block SARS-CoV-2 PLpro activity. A crystal structure of SARS-CoV-2 PLpro in complex with a representative UbV reveals a dimeric UbV bound to PLpro at a site distal to the catalytic site. Yet, the UbV inhibits the essential cleavage activities of the protease in vitro and in cells, and it reduces viral replication in cell culture by almost five orders of magnitude.

SUBMITTER: van Vliet VJE

PROVIDER: S-EPMC9822107 | biostudies-literature | 2022 Dec

REPOSITORIES: biostudies-literature

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Ubiquitin variants potently inhibit SARS-CoV-2 PLpro and viral replication via a novel site distal to the protease active site.

van Vliet Vera J E VJE Huynh Nhan N Palà Judith J Patel Ankoor A Singer Alex A Slater Cole C Chung Jacky J van Huizen Mariska M Teyra Joan J Miersch Shane S Luu Gia-Khanh GK Ye Wei W Sharma Nitin N Ganaie Safder S SS Russell Raquel R Chen Chao C Maynard Mindy M Amarasinghe Gaya K GK Mark Brian L BL Kikkert Marjolein M Sidhu Sachdev S SS

PLoS pathogens 20221222 12

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic has made it clear that combating coronavirus outbreaks benefits from a combination of vaccines and therapeutics. A promising drug target common to all coronaviruses-including SARS-CoV, MERS-CoV, and SARS-CoV-2-is the papain-like protease (PLpro). PLpro cleaves part of the viral replicase polyproteins into non-structural protein subunits, which are essential to the viral replication cycle. Additionally, PLpro can cleave bot ...[more]

PMID: 36548304

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Project description:Protein SUMOylation represents an important cellular process that regulates the activities of numerous host proteins as well as of many invasive viral proteins. Foot-and-mouth disease virus (FMDV) is the first animal virus discovered. However, whether SUMOylation takes place during FMDV infection and what role it plays in FMDV pathogenesis have not been investigated. In the present study, we demonstrated that SUMOylation suppressed FMDV replication by small interfering RNA (siRNA) transfection coupled with pharmaceutical inhibition of SUMOylation, which was further confirmed by increased virus replication for SUMOylation-deficient FMDV with mutations in 3C protease, a target of SUMOylation. Moreover, we provided evidence that four lysine residues, Lys-51, -54, -110, and -159, worked together to confer the SUMOylation to the FMDV 3C protease, which may make SUMOylation of FMDV 3C more stable and improve the host's chance of suppressing the replication of FMDV. This is the first report that four lysine residues can be alternatively modified by SUMOylation. Finally, we showed that SUMOylation attenuated the cleavage ability, the inhibitory effect of the interferon signaling pathway, and the protein stability of FMDV 3C, which appeared to correlate with a decrease in FMDV replication. Taken together, the results of our experiments describe a novel cellular regulatory event that significantly restricts FMDV replication through the SUMOylation of 3C protease. IMPORTANCE FMD is a highly contagious and economically important disease in cloven-hoofed animals. SUMOylation, the covalent linkage of a small ubiquitin-like protein to a variety of substrate proteins, has emerged as an important posttranslational modification that plays multiple roles in diverse biological processes. In this study, four lysine residues of FMDV 3C were found to be alternatively modified by SUMOylation. In addition, we demonstrated that SUMOylation attenuated FMDV 3C function through multiple mechanisms, including cleavage ability, the inhibitory effect of the interferon signaling pathway, and protein stability, which, in turn, resulted in a decrease of FMDV replication. Our findings indicate that SUMOylation of FMDV 3C serves as a host cell defense against FMDV replication. Further understanding of the cellular and molecular mechanisms driving this process should offer novel insights to design an effective strategy to control the dissemination of FMDV in animals.

Dataset Information

Ubiquitin variants potently inhibit SARS-CoV-2 PLpro and viral replication via a novel site distal to the protease active site.

Publications

Ubiquitin variants potently inhibit SARS-CoV-2 PLpro and viral replication via a novel site distal to the protease active site.

Similar Datasets

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