A Viperin Mutant Bearing the K358R Substitution Lost its Anti-ZIKA Virus Activity.

Bénédicte Vanwalscappel,Philippe Desprès,Gilles Gadea

doi:10.3390/ijms20071574

Abstract

Interferon-induced viperin (VP) was identified as playing an important role in the innate immune response against Zika virus (ZIKV). The 361 amino acid long human VP protein comprises of a highly conserved C-terminal region, which has been associated with VP antiviral properties against ZIKV. In the present study, we sought to determine whether the very last C-terminal amino-acid residues of VP might play a role in VP-mediated ZIKV inhibition. To address this issue, a recombinant human viperin (rVPwt) was overexpressed by transfection in human epithelial A549 cells. We confirmed that transient overexpression of rVPwt prior to ZIKV infection dramatically reduced viral replication in A549 cells. Deletion of the last 17 C-terminal amino acids of VP resulted in a higher expression level of mutant protein compared to wild-type VP. Mutational analysis revealed that residue substitution at positions 356 to 360 with five alanine led to the same phenotype. The charged residues Asp356, Lys358, and Asp360 were then identified to play a role in the weak level of VPwt protein in A549 cells. Mutant VP bearing the D360A substitution partially rescued ZIKV growth in A549 cells. Remarkably, a single Lys-to-Arg substitution at position 358 was sufficient to abrogate VP antiviral activity against ZIKV. In conclusion, our study showed that Asp356, Lys358, and Asp360 may have an influence on biochemical properties of VP. Our major finding was that Lys358 was a key amino-acid in VP antiviral properties against ZIKV.

Highlights

The recent emergence of mosquito-borne Zika virus (ZIKV) has been associated with an increase in infection severity in humans, drawing attention to Zika illness as a public health concern worldwide [1]
We investigated the role of amino-acid residues 356 to 360 in VP antiviral properties against ZIKV in human epithelial A549 cells
It has recently been reported that viral clone BR15 derived from a Brazilian epidemic ZIKV strain BeH819015 replicated efficiently in human epithelial A549 cells [8]

Summary

Introduction

The recent emergence of mosquito-borne Zika virus (ZIKV) has been associated with an increase in infection severity in humans, drawing attention to Zika illness as a public health concern worldwide [1]. Viperin (for Virus inhibitory protein endoplasmic reticulum-associated interferon-inducible) has been identified as a major ISG since its expression is induced in response to Type I interferons (IFNs), as well as viral infection [9,10]. In an effort to better understand the molecular basis of VP antiviral action against flaviviruses, it has been observed that VP colocalizes with viral replication complexes, which regroup the NS proteins such as serine protease-NTPase/helicase NS3 [20,21]. Panayiotou et al demonstrated that VP could interact with the NS3 proteins from JEV, TBEV, YFV, and ZIKV and that the VP-mediated degradation of NS3 might account for the restriction of ZIKV replication [13]. VP can catalyze the conversion of cytidine triphosphate (CTP) into 3 -deoxy-3 ,4 -didehydro-CTP (ddhCTP) that acts as a chain terminator for the RNA-dependent RNA polymerase NS5, leading to an arrest in viral RNA replication [22]

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Conclusion