Membrane traffic pathways in viral replication and pathogenesis

Lead Research Organisation: MRC Cell Biology Unit

Abstract

We study the human immunodeficiency virus (HIV) that is linked to AIDS and is believed to have infected 60 million people worldwide. During infection these viruses must interact with receptor proteins expressed on the surfaces of target cells. Using the natural binding proteins of these receptors, it is possible to abrogate HIV infection. We are trying to understand the cellular and molecular mechanisms that underlie this regulation with the aim of developing novel strategies to down regulate the receptors. Such strategies may have efficacy against HIV. Importantly, the receptors exploited by HIV are representative of a large class of similar proteins, so-called G protein coupled receptors (GPCRs), that control a range of biological functions including the immune system. So what we learn about HIV receptors may have relevance to other important biological events.||In addition to understanding virus entry, we also investigate the molecular and cellular mechanisms involved in the formation of new virus particles within infected cells. This requires that the key structural proteins that make up virus particles are bought together at the same time and place within an infected cell so that virus particles can form. We have identified molecular signals in the envelope glycoprotein that are required for the correct trafficking within infected cells. When one of these signals is mutated in a simian model system, the virus is attenuated and is no longer pathogenic. Moreover, animals treated with this virus are protected against challenge with normal pathogenic viruses. Together these studies are providing novel insights to the mechanisms of pathogenesis and may give hints to the key components of immune control.

Technical Summary

We focus on understanding; (i) the mechanisms controlling cell surface expression of cellular receptors for HIV and (ii) the mechanisms involved in the assembly of HIV virions.||Project 1: Human immunodeficiency virus (HIV), the etiological agent of AIDS, infects cells by membrane fusion following interaction with receptors expressed on surfaces of target cells. Two different cell surface glycoproteins form the functional virus receptor - CD4 and a member of the chemokine receptor family of G protein-coupled receptors (GPCRs). Two chemokine receptors have been linked to HIV infection, CC chemokine receptor 5 (CCR5) and CXC chemokine receptor 4 (CXCR4). The agonists for these receptors can protect cells from infection by inducing their endocytosis. We are studying the mechanism of chemokine receptor internalisation and the fate of the internalised molecules. Although focused on chemokine receptors, these studies have relevance for understanding the regulation of a variety of other GPCRs.||Project 2: To make new HIV particles, the essential proteins of the virus must be bought together at the same time and place within infected cells. Structurally, HIV is a relatively simple virus and requires just three virally encoded proteins to make new infectious particles, namely Envelope (Env), Gag and Gag-Pol. We are trying to understand how these proteins are sorted and traffic within cells in order to produce viral particles. We have found that in primary human macrophages, infected with HIV in tissue culture, the assembly events occur on the membranes of a novel intracellular multivesicular compartment. This internal assembly may convey distinct biochemical and functional properties on virus particles and enable release of infectious virus to be regulated and coordinated with the presence of appropriate target cells, such as CD4 positive T cells. Experiments are in progress to determine how the viral components target this compartment and the normal biological functions of this compartment. Significantly, we have found that the Env protein contains conserved signals that control its intracellular trafficking, in part these signals mediate endocytosis and keep the level of Env expressed at the cell surface low. Significantly, in a monkey model, mutation of these signals in the closely related simian immunodeficiency viruses (SIV) leads to attenuation of the pathological potential of the virus. Moreover, animals treated with such viruses show protection when challenged with pathogenic viruses. We hypothesis that correct signals in the structural components of the virus are required to allow efficient assembly of infectious virions and that in animals challenged with viruses lacking the correct signals in the viral structural proteins, the immune system is able to control infection and protect against challenge with non-attenuated viruses.

Publications

10 25 50
 
Description EC FP7 funding
Amount £240,000 (GBP)
Organisation European Commission 
Department Seventh Framework Programme (FP7)
Sector Public
Country European Union (EU)
Start  
 
Description MRC Strategic Grant
Amount £2,000,000 (GBP)
Organisation Medical Research Council (MRC) 
Sector Public
Country United Kingdom
Start 09/2009 
End 09/2012
 
Description UCL MRC Confidence in Concepts
Amount £99,951 (GBP)
Funding ID MC_PC_12024 
Organisation Medical Research Council (MRC) 
Sector Public
Country United Kingdom
Start 10/2013 
End 09/2014
 
Title 1B5, 2C2 
Description Two monoclonal antibody reagents for primate lysosomal membrane proteins. These antibodies are used as morphological markers of the endocytic pathway in studies of HIV replication, dementia, growth factor receptor trafficking, etc. 
Type Of Material Antibody 
Year Produced 2006 
Provided To Others? Yes  
Impact Useful in a variety of cell biological applications - further uses remain to be established. 
 
Description Env trafficking signals in SIV pathogenesis 
Organisation University of Pennsylvania
Department Perelman School of Medicine
Country United States 
Sector Academic/University 
PI Contribution Cell biological analysis of the role of trafficking signals in the cytoplasmic domain of the simian immunodeficiency virus envelope protein - a model for understanding HIV pathogenesis and anti-viral immune responses.
Collaborator Contribution Virological and in vivo studies that complement the cell biology.
Impact Nine publications Multi-disciplinary: Virology, cell biology, in vivo animal models
 
Description HIV-ACE 
Organisation Cochin Institute
Country France 
Sector Academic/University 
PI Contribution EU funded project to identify inhibitor of HIV assembly. We contribute cell biological expertise and SIV model.
Collaborator Contribution Partners provide access to experiment data and novel reagents.
Impact PMID:19864625
Start Year 2008
 
Description Infectious Synapse and Transmission of Disease 
Organisation Julich Research Centre
Country Germany 
Sector Academic/University 
PI Contribution We provide cell biological expertise and experimental models
Collaborator Contribution Program provides complementary experimental systems and access to reagentsProgram provides complementary experimental systems and access to reagentsProgram provides complementary experimental systems and access to reagents
Impact PMID:19143628
Start Year 2006
 
Description Infectious Synapse and Transmission of Disease 
Organisation Rockefeller University
Country United States 
Sector Academic/University 
PI Contribution We provide cell biological expertise and experimental models
Collaborator Contribution Program provides complementary experimental systems and access to reagentsProgram provides complementary experimental systems and access to reagentsProgram provides complementary experimental systems and access to reagents
Impact PMID:19143628
Start Year 2006
 
Description Infectious Synapse and Transmission of Disease 
Organisation University of Geneva
Country Switzerland 
Sector Academic/University 
PI Contribution We provide cell biological expertise and experimental models
Collaborator Contribution Program provides complementary experimental systems and access to reagentsProgram provides complementary experimental systems and access to reagentsProgram provides complementary experimental systems and access to reagents
Impact PMID:19143628
Start Year 2006
 
Description Mode of action of HIV replication inhibitor: CADA 
Organisation University of Leuven
Country Belgium 
Sector Academic/University 
PI Contribution My lab contributed to understanding that CADA can specifically down regulate human CD4 by inhibiting the synthesis of the molecule.
Collaborator Contribution The partners were able to show that CADA can reversibly bind the signal peptide of the nascent human CD4 within the ER translocon and thereby inhibit synthesis of new protein.
Impact A paper is in press in PLOS Biology, other abstracts have been published in meeting reports.
Start Year 2010
 
Description Regulation of GPCR signalling through endocytic trafficking 
Organisation University of California, San Francisco
Country United States 
Sector Academic/University 
PI Contribution My group has provided conceptual input to the work, as well as electron microscopy, data analysis and contributions to the writing of research papers.
Collaborator Contribution The partners have provided conceptual input to the work, as well as live cell imaging, data analysis and contributions to the writing of research papers.
Impact Two papers have been published to date.
Start Year 2010
 
Description Dartford Grammar School Science College, Inaugural Lecture 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Schools
Results and Impact Inaugural lecture for new Science College

Increase awareness of MRC activities, Cell Biology Units work
Year(s) Of Engagement Activity 2009
 
Description MRC Public Panel presentation 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? Yes
Geographic Reach Regional
Primary Audience Public/other audiences
Results and Impact Members of MRC Public Panel came to CBU/LMCB for afternoon to hear talks from myself, other CBU group leaders and MRC Chief Operating Officer (John Jeans)

Positive feedback from panel members
Year(s) Of Engagement Activity 2010
 
Description MRC Showcase on Infection and Immunity 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Health professionals
Results and Impact Poster presentation by group members (A. Pelchen-Matthews, M. Deneka) at MRC Showcase on Infection and Immunity

Improved awareness of team's work
Year(s) Of Engagement Activity 2007
 
Description Naked Scientist/Radio Cambridgeshire 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Public/other audiences
Results and Impact Participation in weekly radio science program - the Naked Scientists - BBC Radio Cambridgeshire and Podcast. Radio show has audience of ~50,000.

Positive feedback from listeners
Year(s) Of Engagement Activity 2010
 
Description Open day/Science Fair 
Form Of Engagement Activity Participation in an open day or visit at my research institution
Part Of Official Scheme? No
Type Of Presentation Workshop Facilitator
Geographic Reach Regional
Primary Audience Public/other audiences
Results and Impact An open day/science fair for MRC funded researchers at UCL - LMCB a principle host and organiser. 500 members of public attended and took part in lectures, demonstrations and other events. Part of MRC Centenary celebrations.

Significant feedback from public impressed by presentations and visibility of MRC funded research.
Year(s) Of Engagement Activity 2013
URL http://www.ucl.ac.uk/lmcb/lmcb-takes-lead-mrc-centenary-celebrations-ucl