ABSTRACT
The establishment of a virus infection is the result of the pathogen's ability to replicate in a hostile environment generated by the host's immune system. Here, we found that ISG15 restricts Dengue and Zika viruses' replication through the stabilization of its binding partner USP18. ISG15 expression was necessary to control DV replication driven by both autocrine and paracrine type one interferon (IFN-I) signaling. Moreover, USP18 competes with NS5-mediated STAT2 degradation, a major mechanism for establishment of flavivirus infection. Strikingly, reconstitution of USP18 in ISG15-deficient cells was sufficient to restore the STAT2's stability and restrict virus growth, suggesting that the IFNAR-mediated ISG15 activity is also antiviral. Our results add a novel layer of complexity in the virus/host interaction interface and suggest that NS5 has a narrow window of opportunity to degrade STAT2, therefore suppressing host's IFN-I mediated response and promoting virus replication.
Subject(s)
Dengue , Interferon Type I , Zika Virus Infection , Zika Virus , Humans , Interferon Type I/metabolism , Zika Virus Infection/genetics , Virus Replication , Dengue/genetics , Ubiquitins/metabolism , Cytokines/metabolism , Ubiquitin Thiolesterase/metabolism , STAT2 Transcription Factor/genetics , STAT2 Transcription Factor/metabolismABSTRACT
Viruses from the Flaviviridae family, such as Dengue virus (DENV), Yellow fever virus (YFV), and Zika virus (ZIKV) are notorious global public health problems. ZIKV emergence in Polynesia and the Americas from 2013 to 2016 raised concerns as new distinguishing features set it apart from previous outbreaks, including its association with neurological complications and heightened disease severity. Virus detection is impaired as cross-reactivity to other closely related orthoflaviviruses is common among commercially available diagnostic kits. While non-structural protein 1 (NS1) has been used as an early marker of DENV and West Nile virus (WNV) infection, little is known about NS1 expression during ZIKV infection. In the present work, we developed a NS1 capture ELISA using a novel ZIKV-specific monoclonal antibody to study NS1 expression dynamics in vitro in mosquito and human cell lines. While detectable in culture supernatants, higher concentrations of NS1 were predominantly cell-associated. To our knowledge, this is the first report of NS1 detection in human cells despite viral clearance over time. Tests with human samples need to be conducted to validate the applicability of NS1 detection for diagnosis, but overall, the tools developed in this work are promising for specific detection of acute ZIKV infection.
Subject(s)
Dengue Virus , Dengue , West Nile Fever , Zika Virus Infection , Zika Virus , Animals , Humans , Antibodies, Viral , Viral Nonstructural Proteins , Enzyme-Linked Immunosorbent Assay , Antibodies, Monoclonal , Sensitivity and SpecificityABSTRACT
IMPORTANCE: One of the fundamental features that make viruses intracellular parasites is the necessity to use cellular translational machinery. Hence, this is a crucial checkpoint for controlling infections. Here, we show that dengue and Zika viruses, responsible for nearly 400 million infections every year worldwide, explore such control for optimal replication. Using immunocompetent cells, we demonstrate that arrest of protein translations happens after sensing of dsRNA and that the information required to avoid this blocking is contained in viral 5'-UTR. Our work, therefore, suggests that the non-canonical translation described for these viruses is engaged when the intracellular stress response is activated.
Subject(s)
Dengue Virus , Stress, Physiological , Virus Replication , Zika Virus , eIF-2 Kinase , Animals , Humans , A549 Cells , Chlorocebus aethiops , Dengue/immunology , Dengue/virology , Dengue Virus/physiology , eIF-2 Kinase/genetics , eIF-2 Kinase/metabolism , Eukaryotic Initiation Factor-2/metabolism , Gene Deletion , Protein Biosynthesis/genetics , Protein Biosynthesis/immunology , Stress, Physiological/genetics , Stress, Physiological/immunology , Vero Cells , Virus Replication/genetics , Virus Replication/immunology , Zika Virus/physiology , Zika Virus Infection/immunology , Zika Virus Infection/virology , RNA, Double-Stranded/metabolismABSTRACT
Yellow fever is an arboviral disease that causes thousands of deaths every year in Africa and the Americas. However, few commercial diagnostic kits are available. Non-structural protein 1 (NS1) is an early marker of several flavivirus infections and is widely used to diagnose dengue virus (DENV) infection. Nonetheless, little is known about the dynamics of Yellow fever virus (YFV) NS1 expression and secretion, to encourage its use in diagnosis. To tackle this issue, we developed a quantitative NS1-capture ELISA specific for YFV using a monoclonal antibody and recombinant NS1 protein. This test was used to quantify NS1 in mosquito and human cell line cultures infected with vaccine and wild YFV strains. Our results showed that NS1 was detectable in the culture supernatants of both cell lines; however, a higher concentration was maintained as cell-associated rather than secreted into the extracellular milieu. A panel of 73 human samples was used to demonstrate the suitability of YFV NS1 as a diagnostic tool, resulting in 80% sensitivity, 100% specificity, a 100% positive predictive value and a 95.5% negative predictive value compared with RT-PCR. Overall, the developed NS1-capture ELISA showed potential as a promising assay for the detection of early YF infection.
