title: A novel interaction between dengue virus nonstructural protein 1 and the NS4A-2K-4B precursor is required for viral RNA replication but not for formation of the membranous replication organelle
creator: Płaszczyca, Anna
creator: Scaturro, Pietro
creator: Neufeldt, Christopher John
creator: Cortese, Mirko
creator: Cerikan, Berati
creator: Ferla, Salvatore
creator: Brancale, Andrea
creator: Pichlmair, Andreas
creator: Bartenschlager, Ralf
subject: ddc-500
subject: 500 Natural sciences and mathematics
subject: ddc-570
subject: 570 Life sciences
description: Dengue virus (DENV) has emerged as major human pathogen. Despite the serious socio-economic impact of DENV-associated diseases, antiviral therapy is missing. DENV replicates in the cytoplasm of infected cells and induces a membranous replication organelle, formed by invaginations of the endoplasmic reticulum membrane and designated vesicle packets (VPs). Nonstructural protein 1 (NS1) of DENV is a multifunctional protein. It is secreted from cells to counteract antiviral immune responses, but also critically contributes to the severe clinical manifestations of dengue. In addition, NS1 is indispensable for viral RNA replication, but the underlying molecular mechanism remains elusive. In this study, we employed a combination of genetic, biochemical and imaging approaches to dissect the determinants in NS1 contributing to its various functions in the viral replication cycle. Several important observations were made. First, we identified a cluster of amino acid residues in the exposed region of the β-ladder domain of NS1 that are essential for NS1 secretion. Second, we revealed a novel interaction of NS1 with the NS4A-2K-4B cleavage intermediate, but not with mature NS4A or NS4B. This interaction is required for RNA replication, with two residues within the connector region of the NS1 "Wing" domain being crucial for binding of the NS4A-2K-4B precursor. By using a polyprotein expression system allowing the formation of VPs in the absence of viral RNA replication, we show that the NS1 -NS4A-2K-4B interaction is not required for VP formation, arguing that the association between these two proteins plays a more direct role in the RNA amplification process. Third, through analysis of polyproteins containing deletions in NS1, and employing a trans-complementation assay, we show that both cis and trans acting elements within NS1 contribute to VP formation, with the capability of NS1 mutants to form VPs correlating with their capability to support RNA replication. In conclusion, these results reveal a direct role of NS1 in VP formation that is independent from RNA replication, and argue for a critical function of a previously unrecognized NS4A-2K-NS4B precursor specifically interacting with NS1 and promoting viral RNA replication.
publisher: PLoS
date: 2019
type: Article
type: info:eu-repo/semantics/article
type: NonPeerReviewed
format: application/pdf
identifier: https://archiv.ub.uni-heidelberg.de/volltextserverhttps://archiv.ub.uni-heidelberg.de/volltextserver/26795/1/Plaszczyca_PPath.pdf
identifier: DOI:10.11588/heidok.00026795
identifier: https://doi.org/10.1371/journal.ppat.1007736
identifier: urn:nbn:de:bsz:16-heidok-267957
identifier:   Płaszczyca, Anna ; Scaturro, Pietro ; Neufeldt, Christopher John ; Cortese, Mirko ; Cerikan, Berati ; Ferla, Salvatore ; Brancale, Andrea ; Pichlmair, Andreas ; Bartenschlager, Ralf  (2019) A novel interaction between dengue virus nonstructural protein 1 and the NS4A-2K-4B precursor is required for viral RNA replication but not for formation of the membranous replication organelle.  PLOS Pathogens, 15 (5).  pp. 1-34.  ISSN 1553-7366 (Druck-Ausg.), 1553-7374 (Online-Ausg.)     
relation: https://archiv.ub.uni-heidelberg.de/volltextserver/26795/
rights: info:eu-repo/semantics/openAccess
rights: http://archiv.ub.uni-heidelberg.de/volltextserver/help/license_urhg.html
language: eng