Abstract
Dengue virus (DENV) is an arthropod-borne Flavivirus that can cause a range of symptomatic disease in humans. There are four dengue viruses (DENV 1 to 4) and infection with one DENV only provides transient protection against a heterotypic virus. Second infections are often more severe as the disease is potentiated by antibodies from the first infection through a process known as antibody dependent enhancement (ADE) of infection. Phosphorylation is a major post-translational modification that can have marked effects on a number of processes. To date there has been little information on the phosphorylation changes induced by DENV infection. This study aimed to determine global phosphoproteome changes induced by DENV 2 in U937 cells infected under an ADE protocol. A 2-dimensional electrophoretic approach coupled with a phosphoprotein-specific dye and mass spectroscopic analysis identified 15 statistically significant differentially phosphorylated proteins upon DENV 2 infection. One protein identified as significantly differentially phosphorylated, pyruvate kinase M2 (PKM2) was validated. Treatment with a PKM2 inhibitor modestly reduced levels of infection and viral output, but no change was seen in cellular viral protein levels, suggesting that PKM2 acts on exocytic virus release. While the effect of inhibition of PKM2 was relatively modest, the results highlight the need for a greater understanding of the role of phosphoproteins in DENV infection.
Highlights
Dengue virus (DENV) is an arthropod-borne Flavivirus that can cause a range of symptomatic disease in humans
After 48 h of infection, it was found that in the absence of antibodies, the percentage of infected cells was 16.340 ± 2.286% (Supplementary Fig. 1), while an antibody dilution of 1:200 resulted in the highest percentage of infection of 69.780 ± 0.710% which was significantly different from the direct infection (P < 0.05), confirming the cells were infected under a condition of antibody dependent enhancement (ADE)
To determine the role of pyruvate kinase M2 (PKM2) phosphorylation in DENV 2 infection, we evaluated the effects of a PKM2 inhibitor and activator
Summary
Dengue virus (DENV) is an arthropod-borne Flavivirus that can cause a range of symptomatic disease in humans. An additional study reported that increased DENV disease severity correlated with high viremia titer, secondary DENV infection and DENV 2 virus s erotype[6]. There is an increased number of infected cells due to increased antibody-mediated cell binding and entry of both mature and (partially) immature DENV particles which is known as extrinsic ADE. Another form called intrinsic ADE occurs through increased virus production per infected cell due to suppression of the innate antiviral response[10]. Intrinsic ADE of DENV infection is believed to involve suppression of the toll-like receptor (TLR) and retinoic acid inducible protein I/melanoma differentiation-associated gene[5] (RIG-I/MDA5) signaling pathways thereby decreasing the production of type I interferon and interferonactivated antiviral m olecules[11]
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