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Analysis of Zika virus capsid-Aedes aegypti mosquito interactome reveals pro-viral host factors critical for establishing infection.

ABSTRACT: The escalating global prevalence of arboviral diseases emphasizes the need to improve our understanding of their biology. Research in this area has been hindered by the lack of molecular tools for studying virus-mosquito interactions. Here, we develop an Aedes aegypti cell line which stably expresses Zika virus (ZIKV) capsid proteins in order to study virus-vector protein-protein interactions through quantitative label-free proteomics. We identify 157 interactors and show that eight have potentially pro-viral activity during ZIKV infection in mosquito cells. Notably, silencing of transitional endoplasmic reticulum protein TER94 prevents ZIKV capsid degradation and significantly reduces viral replication. Similar results are observed if the TER94 ortholog (VCP) functioning is blocked with inhibitors in human cells. In addition, we show that an E3 ubiquitin-protein ligase, UBR5, mediates the interaction between TER94 and ZIKV capsid. Our study demonstrates a pro-viral function for TER94/VCP during ZIKV infection that is conserved between human and mosquito cells.

SUBMITTER: Gestuveo RJ

PROVIDER: S-EPMC8119459 | biostudies-literature | 2021 May

REPOSITORIES: biostudies-literature

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Analysis of Zika virus capsid-Aedes aegypti mosquito interactome reveals pro-viral host factors critical for establishing infection.

Gestuveo Rommel J RJ Royle Jamie J Donald Claire L CL Lamont Douglas J DJ Hutchinson Edward C EC Merits Andres A Kohl Alain A Varjak Margus M

Nature communications 20210513 1

The escalating global prevalence of arboviral diseases emphasizes the need to improve our understanding of their biology. Research in this area has been hindered by the lack of molecular tools for studying virus-mosquito interactions. Here, we develop an Aedes aegypti cell line which stably expresses Zika virus (ZIKV) capsid proteins in order to study virus-vector protein-protein interactions through quantitative label-free proteomics. We identify 157 interactors and show that eight have potenti ...[more]

PMID: 33986255

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Project description:The yellow fever mosquito, Aedes aegypti, serves as the primary vector for epidemic transmission of yellow fever, dengue, Zika (ZIKV), and chikungunya viruses to humans. Control of Ae. aegypti is currently limited to insecticide applications and larval habitat management; however, to combat growing challenges with insecticide resistance, novel genetic approaches for vector population reduction or transmission interruption are being aggressively pursued. The objectives of this study were to assess the ability of the Ae. aegypti antiviral exogenous-small interfering RNA (exo-siRNA) response to inhibit ZIKV infection and transmission, and to identify the optimal RNA interference (RNAi) target region in the ZIKV genome. We accomplished these objectives by in vitro transcription of five long double-stranded RNAs (dsRNAs) from the genome region spanning the NS2B-NS3-NS4A genes, which were the most highly conserved among ZIKV RNA sequences representing both East and West African and Asian-American clades, and evaluation of the ability of these dsRNAs to trigger an effective antiviral exo-siRNA response after intrathoracic injection into Ae. aegypti. In a pilot study, five ZIKV dsRNAs were tested by intrathoracic inoculation of 250?ng dsRNA into groups of approximately 5-day-old mosquitoes. Three days post-inoculation, mosquitoes were provided an infectious blood-meal containing ZIKV strain PRVABC59 (Puerto Rico), MR766 (Uganda), or 41525 (Senegal). On days 7 and 14 post-infection individual whole mosquito bodies were assessed for ZIKV infectious titer by plaque assays. Based on the results of this initial assessment, three dsRNAs were selected for further evaluation of viral loads of matched body and saliva expectorants using a standardized infectious dose of 1?×?107?PFU/mL of each ZIKV strain. Fourteen days post-exposure to ZIKV, paired saliva and carcass samples were harvested from individual mosquitoes and assessed for ZIKV RNA load by qRT-PCR. Injection of each of the three dsRNAs resulted in significant inhibition of replication of all three strains of ZIKV in mosquito bodies and saliva. This study lays critical groundwork for pursuing ZIKV transmission-blocking strategies that exploit the Ae. aegypti exo-siRNA response for arbovirus suppression in natural populations.

Project description:Biological control of mosquito vectors using the endosymbiotic bacteria Wolbachia is an emerging strategy for the management of human arboviral diseases. We recently described the development of a strain of Aedes aegypti infected with the Wolbachia strain wAlbB (referred to as the wAlbB2-F4 strain) through simple backcrossing of wild type Australian mosquitoes with a wAlbB infected Ae. aegypti strain from the USA. Field releases of male wAlbB2-F4 mosquitoes resulted in the successful suppression of wild populations of mosquitoes in the trial sites by exploiting the strain's Wolbachia-induced cytoplasmic incompatibility. We now demonstrate that the strain is resistant to infection by dengue and Zika viruses and is genetically similar to endemic Queensland populations. There was a fourfold reduction in the proportion of wAlbB2-F4 mosquitoes that became infected following a blood meal containing dengue 2 virus (16.7%) compared to wild type mosquitoes (69.2%) and a 6-7 fold reduction in the proportion of wAlbB2-F4 mosquitoes producing virus in saliva following a blood meal containing an epidemic strain of Zika virus (8.7% in comparison to 58.3% in wild type mosquitoes). Restriction-site Associated DNA (RAD) sequencing revealed that wAlbB2-F4 mosquitoes have > 98% Australian ancestry, confirming the successful introduction of the wAlbB2 infection into the Australian genomic background through backcrossing. Genotypic and phenotypic analyses showed the wAlbB2-F4 strain retains the insecticide susceptible phenotype and genotype of native Australian mosquitoes. We demonstrate that the Wolbachia wAlbB2-F4, in addition to being suitable for population suppression programs, can also be effective in population replacement programs given its inhibition of virus infection in mosquitoes. The ease at which a target mosquito population can be transfected with wAlbB2, while retaining the genotypes and phenotypes of the target population, shows the utility of this strain for controlling the Ae. aegypti mosquitoes and the pathogens they transmit.

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Analysis of Zika virus capsid-Aedes aegypti mosquito interactome reveals pro-viral host factors critical for establishing infection.

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Analysis of Zika virus capsid-Aedes aegypti mosquito interactome reveals pro-viral host factors critical for establishing infection.

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