Abstract
The Japanese encephalitis virus (JEV) is a mosquito-borne flavivirus prevalent in east and southeast Asia, the Western Pacific, and northern Australia. Since viruses are obligatory intracellular pathogens, the dynamic processes of viral entry, replication, and assembly are dependent on numerous host-pathogen interactions. Efforts to identify JEV-interacting host factors are ongoing because their identification and characterization remain incomplete. Three enzymatic activities of flavivirus non-structural protein 3 (NS3), including serine protease, RNA helicase, and triphosphatase, play major roles in the flaviviruses lifecycle. To identify cellular factors that interact with NS3, we screened a human brain cDNA library using a yeast two-hybrid assay, and identified eight proteins that putatively interact with NS3: COPS5, FBLN5, PPP2CB, CRBN, DNAJB6, UBE2N, ZNF350, and GPR137B. We demonstrated that the DnaJ heat shock protein family (Hsp40) member B6 (DNAJB6) colocalizes and interacts with NS3, and has a negative regulatory function in JEV replication. We also show that loss of DNAJB6 function results in significantly increased viral replication, but does not affect viral binding or internalization. Moreover, the time-course of DNAJB6 disruption during JEV infection varies in a viral load-dependent manner, suggesting that JEV targets this host chaperone protein for viral benefit. Deciphering the modes of NS3-interacting host proteins functions in virion production will shed light on JEV pathogenic mechanisms and may also reveal new avenues for antiviral therapeutics.
Highlights
Viruses of the Flaviviridae family are responsible for significant morbidity and mortality, and are increasing in prevalence worldwide
Sequence analysis revealed eight cellular proteins that potentially interact with Japanese encephalitis virus (JEV) non-structural protein 3 (NS3) (COPS5, FBLN5, PPP2CB, CRBN, DNAJB6, UBE2N, ZNF350, and GPR137B; Table 1)
SRciN. 2A019l,e2v0e,l5s71w9ere significantly higher in ΔDNAJB6 cells than in parental HEK293 7 of 18 cells as measured by RT-qPCR (Figure 5C). These results show that the infectivity of JEV in ΔDNAJB6 cells is significantly enhanced over parental cells
Summary
Viruses of the Flaviviridae family are responsible for significant morbidity and mortality, and are increasing in prevalence worldwide. Mosquito-borne diseases caused by flaviviruses have spread rapidly in recent years. The prevalence of Dengue, West Nile, and Japanese encephalitis, three major mosquito-borne viruses, are an expanding public health threat. These pathogens currently threaten the health of millions of individuals, and are a potential threat in non-endemic areas that are colonized by mosquitos [1]. The Japanese encephalitis virus (JEV), a member of the genus Flavivirus, has the potential to become a global pathogen. With the rise in global temperatures, and the increased range of competent mosquitoes, the spread of JEV to Europe and North America appears probable [7]. Since viruses are obligatory intracellular pathogens, an in-depth understanding of the mechanisms of the replication cycle and the interactions with host organisms are required to develop strategies to combat their infection [8]
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