Dataset Information

An equine iPSC-based phenotypic screening platform identifies pro- and anti-viral molecules against West Nile virus.

ABSTRACT: Outbreaks of West Nile virus (WNV) occur periodically, affecting both human and equine populations. There are no vaccines for humans, and those commercialised for horses do not have sufficient coverage. Specific antiviral treatments do not exist. Many drug discovery studies have been conducted, but since rodent or primate cell lines are normally used, results cannot always be transposed to horses. There is thus a need to develop relevant equine cellular models. Here, we used induced pluripotent stem cells to develop a new in vitro model of WNV-infected equine brain cells suitable for microplate assay, and assessed the cytotoxicity and antiviral activity of forty-one chemical compounds. We found that one nucleoside analog, 2'C-methylcytidine, blocked WNV infection in equine brain cells, whereas other compounds were either toxic or ineffective, despite some displaying anti-viral activity in human cell lines. We also revealed an unexpected proviral effect of statins in WNV-infected equine brain cells. Our results thus identify a potential lead for future drug development and underscore the importance of using a tissue- and species-relevant cellular model for assessing the activity of antiviral compounds.

SUBMITTER: Cochet M

PROVIDER: S-EPMC10943879 | biostudies-literature | 2024 Mar

REPOSITORIES: biostudies-literature

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An equine iPSC-based phenotypic screening platform identifies pro- and anti-viral molecules against West Nile virus.

Cochet Marielle M Piumi François F Gorna Kamila K Berry Noémie N Gonzalez Gaëlle G Danckaert Anne A Aulner Nathalie N Blanchet Odile O Zientara Stéphan S Donadeu Francesc Xavier FX Munier-Lehmann Hélène H Richardson Jennifer J Benchoua Alexandra A Coulpier Muriel M

Veterinary research 20240316 1

Outbreaks of West Nile virus (WNV) occur periodically, affecting both human and equine populations. There are no vaccines for humans, and those commercialised for horses do not have sufficient coverage. Specific antiviral treatments do not exist. Many drug discovery studies have been conducted, but since rodent or primate cell lines are normally used, results cannot always be transposed to horses. There is thus a need to develop relevant equine cellular models. Here, we used induced pluripotent ...[more]

PMID: 38493182

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Project description:BackgroundWest Nile virus (WNV) is a single-stranded RNA virus that can cause neurological disease in both humans and horses. Due to the movement of competent vectors and viraemic hosts, WNV has repeatedly emerged globally and more recently in western Europe. Within the UK, WNV is a notifiable disease in horses, and vaccines against the virus are commercially available. However, there has been no investigation into the seroprevalence of WNV in the UK equine population to determine the extent of vaccination or to provide evidence of recent infection.MethodsEquine serum samples were obtained from the Animal and Plant Health Agency's equine testing service between August and November 2019. A total of 988 serum samples were selected for horses resident in South East England. WNV seroprevalence was determined using two enzyme-linked immunosorbent assays (ELISAs) to detect total flavivirus antibodies and WNV-specific immunoglobulin M (IgM) antibodies. Positive IgM results were investigated by contacting the submitting veterinarian to establish the clinical history or evidence of prior vaccination of the horses in question.ResultsWithin the cohort, 274 samples tested positive for flavivirus antibodies, of which two subsequently tested positive for WNV-specific IgM antibodies. The follow-up investigation established that both horses had been vaccinated prior to serum samples being drawn, which resulted in an IgM-positive response. All the samples that tested positive by competition ELISA were from horses set to be exported to countries where WNV is endemic. Consequently, the positive results were likely due to previous vaccination. In contrast, 714 samples were seronegative, indicating that the majority of the UK equine population may be susceptible to WNV infection.ConclusionsThere was no evidence for cryptic WNV infection in a cohort of horses sampled in England in 2019. All IgM-seropositive cases were due to vaccination; this should be noted for future epidemiological surveys in the event of a disease outbreak, as it is not possible to distinguish vaccinated from infected horses without knowledge of their clinical histories.

Project description:Upon infection, viruses remodel host cells to create an environment hospitable to viral replication. In response, host cells have evolved to sense infection and rapidly induce pathways to limit viral replication. One strategy used by both players is to regulate the host cell proteome for their benefit. To characterize these changes, we used a mass spectrometry strategy to define alterations in the viral and host proteome during infection of human cells with West Nile virus (WNV). Our studies reveal previously uncharacterized post-translational modifications (PTMs) on viral proteins, including phosphorylation sites on the WNV nonstructural proteins, NS3 and NS5, that impact viral RNA replication. Additionally, we monitored changes in host protein abundance and found that while 82 proteins had increased expression upon WNV infection, only five of these are canonical interferon stimulated genes (ISGs). Our data suggests that the majority of the upregulated non-ISGs are regulated at the translational, rather than transcriptional level, defining a new group of virus-induced proteins (VIPs). HERPUD1 is a stress-response protein that was induced at both the transcriptional and translational level. We show HERPUD1 restricts WNV replication, likely through a mechanism independent of its previously defined role in ER-associated degradation (ERAD). Our phosphoproteomics studies revealed eight phosphorylation sites on viral proteins, including a site on the WNV helicase/protease NS3 that is important for viral replication. Further, we identified XXX phosphorylation sites on host proteins with increased abundance and XXX sites with decreased abundance during WNV infection. We used bioinformatics to identify phosphosites predicted to increase host protein activity and found two antiviral host kinases that are activated by phosphorylation during WNV infection. First, we identified a phosphorylation site at S108 of AMPKβ1, a non-catalytic subunit that is thought to regulate activity of the AMPK complex. We also found that a group I p21-activated kinase, PAK2, is phosphorylated at S141. This phosphorylation event in the autoinhibitory domain activates PAK2, which restricts viral protein translation. Altogether, our work contributes to our understanding of the interplay between host and virus while providing a resource to define the changes to the proteome that regulate viral infection.

Dataset Information

An equine iPSC-based phenotypic screening platform identifies pro- and anti-viral molecules against West Nile virus.

Publications

An equine iPSC-based phenotypic screening platform identifies pro- and anti-viral molecules against West Nile virus.

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