[Source: Journal of Virology, full page: (LINK). Abstract, edited.]
Probing Zika virus neutralization determinants with glycoprotein mutants bearing linear epitope insertions
Matthew T. Chambers a, Megan C. Schwarz a, Marion Sourisseau a, Essanna S. Gray a and Matthew J. Evans a#
Author Affiliations: a Department of Microbiology, Icahn School of Medicine at Mount Sinai, New York, NY, 10029
ABSTRACT
Zika virus (ZIKV) glycoproteins are the primary target of the humoral immune response. In this study, we explored the capacity of these glycoproteins to tolerate insertion of linear epitope sequences, and the potential of antibodies that bind these epitopes to inhibit infection. We first created a panel of ZIKV mutants with the FLAG epitope inserted in premembrane (prM) and envelope (E) glycoprotein regions. Insertion locations were based on the results of our recent transposon insertional mutagenesis screen. Although FLAG insertions in prM greatly impaired viral fitness, this sequence was tolerated in numerous surface-exposed E protein sites. We observed that mutants bearing FLAG epitopes in E domains I, II, and the DI-II hinge region were all neutralized by FLAG antibody, however, the neutralization sensitivity varied highly. We measured antibody binding efficiency and found that this closely matched the pattern of neutralization sensitivity. We determined that E glycosylation did not affect antibody binding to a nearby epitope or its capacity of to serve as a neutralization target. Although we could not generate infectious viruses with FLAG epitope insertions in a buried region of E protein domain III, we found that the V5 epitope could be inserted without greatly impacting fitness. Furthermore, this virus was efficiently neutralized by V5 antibodies, highlighting that even buried epitopes can function as neutralization targets. Finally, we analyzed the timing of antibody neutralization activity during cell entry and found that all antibodies blocked a step after cell attachment.
IMPORTANCE
Zika virus (ZIKV) infections are associated with severe birth defects and neurological disease. The structure of the mature ZIKV particle reveals a virion surface covered by the envelope glycoprotein, which is the dominant target of the humoral immune response. It is unclear if all regions of the envelope protein surface, or even ‘buried’ epitopes, can function as neutralization targets. To test this, we created a panel of ZIKV mutants with epitope insertions in different regions of the envelope protein. In characterizing these viruses, we found that the strength of antibody binding to an epitope is the major determinant of the neutralization potential of an antibody, even a buried region of the envelope protein can be efficiently targeted, and the sole potential envelope glycan does not impact nearby epitope antibody binding and neutralization. Furthermore, this work provides important insights into our understanding of how antibodies neutralize ZIKV.
FOOTNOTES
#Corresponding author: Matthew J. Evans, Ph.D., Department of Microbiology, Icahn School of Medicine at Mount Sinai, 1 Gustave L. Levy Place, Box 1124, New York, NY 10029, Tel: 212-241-7319, Fax: 212-534-1684, Email: matthew.evans@mssm.edu
Copyright © 2018 American Society for Microbiology. All Rights Reserved.
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Keywords: Zika Virus; Virology; Viral Pathogenesis.
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