Vaccinia virus entry, exit and evasion
Lead Research Organisation:
Imperial College London
Department Name: Dept of Medicine
Abstract
Viruses remain a potent threat to human health as illustrated by the current epidemics caused by influenza virus and human immunodeficiency virus. Basic research on how viruses replicate, spread and evade our immunological defences is important in advancing our understanding of how viruses cause disease and will underpin the development of new vaccines and anti-viral drugs. This proposal concerns vaccinia virus (VACV), a poxvirus and the live vaccine that was used to eradicate smallpox. Although smallpox has been eradicated, VACV continues to be studied because i) it is possible to engineer VACV as a vaccine against other infectious diseases, and ii) the interaction between VACV and the host cell and immune system is providing novel, fundamental information about how viruses cause disease. This application will focus on how VACV enters cells, how new virus particles are transported out of the cell, how these virus particles spread rapidly to other cells, and how VACV evades the immune response to infection. The information obtained will facilitate the engineering of VACV as a safer and more immunogenic vaccine for infectious diseases and cancer.
Technical Summary
Vaccinia virus (VACV) is the live vaccine that was used to eradicate smallpox. Thereafter, VACV has continued to be studied due to its potential as a live recombinant vaccine for other diseases, and because it is an excellent model for studying virus-host interactions and immune evasion. This project proposal is for renewal of an existing programme grant and the objectives are to study fundamental aspects of VACV biology including entry, egress, spread and immune evasion.
1. Entry. Studies of VACV entry into cells will focus on the double enveloped extracellular virion. Specifically, we will study how the entry of this virus into cells by a non-fusogenic shedding of the outer envelope is prevented when this virion contacts the A33/A36 complex on the infected cell surface.
2. Intracellular transport. We will investigate how newly formed VACV particles are transported on microtubules to the cell surface and the roles of proteins E2, 12 and A36 in this process. This work will build on our recent demonstration that F12 is a structural mimic of kinesin light chain (part of the kinesin motor) and proteins E2, F12 and A36 all possess tetratricopeptide repeats that mediate protein:protein interactions.
3. Virus spread. Recently we have discovered a novel mechanism that VACV uses to spread from cell to cell at a rate 4 times faster than can be explained by the virus replication kinetics. The mechanism involves repulsion of superinfecting virions by the A33 / A36 protein complex so that virus particles are disseminated to distant uninfected cells without replication being required in the intervening cells. This study has redefined how viruses form a plaque and reveals another target for intervention with anti-viral therapy. We will investigate which parts of these proteins are needed, which proteins on the virus surface are required, and test whether soluble fragments of either can diminish virus spread.
4. Lastly we will investigate immune evasion by VACV, with focus on i) VACV encoded miRNAs that are produced from within the coding region of a VACV mRNA, and ii) an uncharacterised protein, B6, that we hypothesise is a novel chemokine binding protein. The contribution of each to virus virulence and immune evasion will be investigated using appropriate in vitro and in vivo models.
1. Entry. Studies of VACV entry into cells will focus on the double enveloped extracellular virion. Specifically, we will study how the entry of this virus into cells by a non-fusogenic shedding of the outer envelope is prevented when this virion contacts the A33/A36 complex on the infected cell surface.
2. Intracellular transport. We will investigate how newly formed VACV particles are transported on microtubules to the cell surface and the roles of proteins E2, 12 and A36 in this process. This work will build on our recent demonstration that F12 is a structural mimic of kinesin light chain (part of the kinesin motor) and proteins E2, F12 and A36 all possess tetratricopeptide repeats that mediate protein:protein interactions.
3. Virus spread. Recently we have discovered a novel mechanism that VACV uses to spread from cell to cell at a rate 4 times faster than can be explained by the virus replication kinetics. The mechanism involves repulsion of superinfecting virions by the A33 / A36 protein complex so that virus particles are disseminated to distant uninfected cells without replication being required in the intervening cells. This study has redefined how viruses form a plaque and reveals another target for intervention with anti-viral therapy. We will investigate which parts of these proteins are needed, which proteins on the virus surface are required, and test whether soluble fragments of either can diminish virus spread.
4. Lastly we will investigate immune evasion by VACV, with focus on i) VACV encoded miRNAs that are produced from within the coding region of a VACV mRNA, and ii) an uncharacterised protein, B6, that we hypothesise is a novel chemokine binding protein. The contribution of each to virus virulence and immune evasion will be investigated using appropriate in vitro and in vivo models.
