Dengue Virus Hijacks a Noncanonical Oxidoreductase Function of a Cellular Oligosaccharyltransferase Complex

Abstract

Dengue virus (DENV) is the most common arboviral infection globally, infecting an estimated 390 million people each year. We employed a genome-wide clustered regularly interspaced short palindromic repeat (CRISPR) screen to identify host dependency factors required for DENV propagation and identified the oligosaccharyltransferase (OST) complex as an essential host factor for DENV infection. Mammalian cells express two OSTs containing either STT3A or STT3B. We found that the canonical catalytic function of the OSTs as oligosaccharyltransferases is not necessary for DENV infection, as cells expressing catalytically inactive STT3A or STT3B are able to support DENV propagation. However, the OST subunit MAGT1, which associates with STT3B, is also required for DENV propagation. MAGT1 expression requires STT3B, and a catalytically inactive STT3B also rescues MAGT1 expression, supporting the hypothesis that STT3B serves to stabilize MAGT1 in the context of DENV infection. We found that the oxidoreductase CXXC active site motif of MAGT1 was necessary for DENV propagation, as cells expressing an AXXA MAGT1 mutant were unable to support DENV infection. Interestingly, cells expressing single-cysteine CXXA or AXXC mutants of MAGT1 were able to support DENV propagation. Utilizing the engineered peroxidase APEX2, we demonstrate the close proximity between MAGT1 and NS1 or NS4B during DENV infection. These results reveal that the oxidoreductase activity of the STT3B-containing OST is necessary for DENV infection, which may guide the development of antiviral agents targeting DENV.

Single-cell transcriptional dynamics of flavivirus infection

“…Several high-throughput approaches have been applied to screen all 20,000 human genes for interactions with flaviviruses, including knockdown screens based on RNA interference (Sessions et al 2009;Kwon et al 2014;Le Sommer et al 2012) , knockout screens via haploid cell lines or CRISPR libraries (Marceau et al 2016;Lin et al 2017) , and bulk transcriptomics via microarrays or RNA-Seq (Sessions et al 2013;Moreno-Altamirano et al 2004;Fink et al 2007;Conceição et al 2010;Becerra et al 2009;Liew and Chow 2006) . While these approaches have provided important insights, our understanding of infection-triggered cellular responses is far from complete.…”

Targeting STT3A‐oligosaccharyltransferase with NGI‐1 causes herpes simplex virus 1 dysfunction

“…OST catalytic subunits, therefore, support HSV-1 replication (and are the targets of NGI-1) by providing an important catalytic function, not a noncatalytic structural role. This contrasted with the finding that noncatalytic structural features of OST catalytic subunits were critical for dengue virus replication (26,27). Glycosylation indices for gC ( Figs.…”

“…Work with flaviviruses showed that although both STT3A-OST and STT3B-OST were necessary for dengue virus infection, these enzyme complexes acted as structural scaffolds for replicating viral components rather than as catalytic OSTs per se (26,27,29). This counterintuitive result was echoed by showing that NGI-1 impaired dengue virus replication by interaction with OST independent of its enzymatic activity (29).…”

“…Replication of several pathogenic mosquito-borne flaviviruses (dengue, West Nile, and yellow fever) is suppressed by loss of STT3A, as well as by loss of STT3B in the case of dengue virus (26)(27)(28). Infection by these viruses is also inhibited by NGI-1 in the concentration range that impairs N-glycosylation of their envelope glycoproteins.…”

“…Interestingly, although OST complexes are the targets of NGI-1, NGI-1's ability to inhibit dengue virus replication is not directly linked to its inhibition of actual OST enzymatic activity (29). Moreover, OST complexes with catalytic subunit mutations still support dengue virus replication (26,27). This indicates an unanticipated complexity of the effects of OST manipulation on viral replication.…”