UK - Department for Business, Energy and Industrial Strategy
Analysis of flavivirus infection on the cellular lipidome - implications for virus particle production and replication.
Project Data Last Updated: 27/08/2020
IATI Identifier: GB-GOV-13-FUND--Newton-MR_R020566_1
Flaviviruses are arthropod-borne RNA viruses that cause significant human morbidity and mortality worldwide. Over the last 50 years, a number of flaviviruses including; dengue virus (DENV), Japanese encephalitis virus (JEV), West Nile virus (WNV) and recently Zika virus (ZIKV) have emerged to cause diseases that are now serious global public health concerns. Countries in tropical and sub-tropical regions, such as Thailand, bear the greatest economic and societal cost of these diseases, which are endemic due to the presence of the mosquito vectors. It is estimated that DENV infections alone cost Thailand some US$290 million per year in direct and indirect costs. However, fully protective and safe vaccines for prevention of mosquito-borne flavivirus diseases are only available for yellow fever virus (YFV) and JEV. As such, there is a need to develop improved flavivirus vaccines and anti-viral therapies. A greater understanding of the interaction between flaviviruses and their hosts will facilitate improved control measures. Previous studies have established that flaviviruses modulate cellular lipid metabolism to promote virus replication and assembly and that changes in serum lipids are associated with disease severity. However, there is little known concerning how the overall changes to lipid metabolism during infection specifically affect i) the lipid composition of the virus particle and its infectivity ii) the types of lipids that are secreted from infected cells and iii) disease processes. Little is also known concerning how different flaviviruses alter the host and virus lipidome and whether common lipids are involved. The UK and Thai investigators have already conducted preliminary studies using high-throughput mass spectrometry (MS) to investigate how DENV infection i) modulates intracellular and secreted protein profiles ii) modulates the cellular lipidome and iii) influences the composition of DENV particles. The studies have revealed that proteins involved in lipid metabolism and/or lipoproteins are differentially regulated and secreted during DENV infection and that the lipidomic profile of DENV-2 infected cells is altered, compared to uninfected cells. In this investigation we propose to use advanced high-throughput liquid chromatography-MS to analyse how lipid metabolism is perturbed during flavivirus infection and specifically define the lipids that are key for flavivirus replication and particle assembly. Specifically, we will examine the lipid content of infectious flavivirus particles and virus-like-particles (VLPs) and the cellular membranes used for flavivirus replication during infection. We will also determine whether flavivirus infection results in changes in the lipids secreted from cells. The studies will be done using three flaviviruses of concern in Thailand; DENV, JEV and ZIKV, to determine if flaviviruses rely on common changes to lipid metabolism for their replication and particle assembly. The importance of these lipids in infection will be confirmed by altering their synthesis by RNA knockdown and/or drug treatment. We will also determine whether the lipid composition of virus particles affects the host response to viral entry and the immunogenicity of virus particles. In the longer term, this knowledge will be used as a basis to i) manipulate cellular lipid synthesis to increase the secretion of VLPs to produce improved vaccines ii) identify lipid biosynthetic pathways essential for flavivirus replication and potentially targets for novel anti-viral therapies and iii) increase our knowledge of viral pathogenesis. The project will build on existing collaborations between the UK and Thai partners and develop the capability of the Thai partners to use high-throughput approaches to analyse virus particles and how viral infection remodels the cellular lipidome, not only for flaviviruses, but also for other emerging arboviruses for which no therapies exist.Objectives
Flaviviruses such as dengue virus (DENV), Japanese encephalitis virus (JEV) and Zika virus (ZIKV) are arthropod-borne RNA viruses that have emerged in recent years to become serious global public health concerns. Countries in tropical and sub-tropical regions, such as Thailand, bear the greatest economic and societal cost of these diseases, which are endemic due to the presence of the mosquito vectors. Flaviviruses are known to manipulate cellular lipid metabolism during infection to promote viral replication, particle assembly and egress which influences virus infectivity and disease. We hypothesize that the alterations that occur to the lipid composition of the host cell membranes used for flavivirus replication and assembly define the lipid composition and infectivity of virus particles and modulate the lipidomic secretome of infected cells. The OVERALL AIM of this project is answer the following scientific questions: 1) Are the changes that occur in the composition of the host cell membranes used for flavivirus replication and assembly reflected in the lipid composition of the flavivirus particle and the lipids that are secreted from cells? 2) Are the specific lipids present in flavivirus particles important for the production and immunogenic properties of either flavivirus particles or empty virus-like- particles (VLPs), that have potential as vaccines? 3) Are flaviviruses reliant on common lipids for their replication? In order to achieve our overall aim the MAIN OBJECTIVES of the project are to: 1) Use a high-throughput liquid chromatography (LC) mass spectrometry (MS) approach to: a) Investigate whether the lipid profiles of intracellular membranes remodelled during flavivirus (DENV, JEV and ZIKV) replication reflect the lipid composition of flavivirus particles. b) Analyse the alterations that occur in the cellular lipidomic secretome during flavivirus infection. c) Determine whether there are differences in the lipid composition of flavivirus VLPs produced from cells containing an active flavivirus replication complex compared to uninfected cells. 2) Use complementary approaches to confirm that the lipids identified to be associated with virus replication and/or particle morphogenesis by LC-MS analysis are essential for productive virus infection. 3) Determine whether the lipid composition of VLPs affects VLP yields, immunogenicity or intracellular signalling. The collaborative scientific investigation will enhance the capacity of Thai researchers to undertake advanced LC-MS based lipidomic analysis through academic exchange and training. The approach used in this investigation can be applied to other investigations on infectious diseases. The research will improve our understanding of the role of lipids in the flavivirus particle that will have direct benefits to controlling these viruses through improved VLP based vaccine production and the identification of targets for anti-viral therapy which are both active fields of research Thailand.
|Extending:||UK Research & Innovation|
|Funding:||UK - Department for Business, Energy and Industrial Strategy|
|Implementing:||University of Bristol|
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