Insect Virus Research Group

Professor Linda King, Dr Caroline Griffiths

 

The insect virus research group is housed in a purpose built suite of laboratories which are well equipped for many aspects of molecular biology and virology. The group also utilises departmental facilities including a state-of-the-art microscopy facility for confocal and electron microscopy.

We undertake both basic and applied research in the field of insect virology. Within the research group, interests include both fundamental studies on the replication and pathogenesis of insect viruses in cell culture and in the natural caterpillar hosts, and on the exploitation of these viruses as protein expression vectors in insect and mammalian cells. We are particularly interested in the interrelated roles of three virus proteins, cathepsin, chitinase and P10 and the interactions between virus proteins and the host cell cytoskeleton and endomembrane systems. We are also engaged in the exploitation of baculoviruses as expression vectors for the production of proteins for use in biotechnology, biological research and medicine.

Our aim is to use our understanding of the replication and pathogenesis of the virus to improve the baculovirus expression vector system. After many years of offering an in-house baculovirus expression consultancy service, the group successfully spun out its commercial activities in October 2007 as Oxford Expression Technologies Ltd.

Projects

Baculovirus transmission and interactions with host larvae: understanding the role of accessory proteins

Viruses are molecular parasites that invade living cells and redirect their biological functions to their own purposes, namely the propagation of new viruses. This parasitic nature of viruses means that they possess very limited genetic information, hijacking many host cell key proteins and enzymes. Nevertheless, many viruses possess more genetic information than the ‘bare minimum’ and baculoviruses are known to encode several proteins that are found to be non-essential for the propagation of new viruses in cells. These non-essential, or auxiliary, proteins are of significant interest and have already been shown to provide unexpected functions that contribute to the replication and transmission of the virus in its natural insect host.
 
Our studies have investigated two auxiliary proteins, chitinase and cathepsin, synthesised by the baculovirus AcMNPV, which work in concert to liquefy the chitinous tissues of the caterpillar hosts in the late stages of infection. By manipulating the genome of this virus to disrupt one, or both, of the genes encoding these proteins, the rate of horizontal transmission of the virus between hosts was reduced when either or both proteins were missing. More recent studies are investigating an auxiliary protein, P10, which appears to influence the integrity of infected cells. This in turn may have also implications for virus release and transmission, key parameters in the survival of viruses in the natural populations.
 
The regulation of baculovirus late gene expression is also being studied using molecular genetics, protein engineering and immunocytochemical techniques. The role of baculovirus genes in the transmission of virus is also being examined together with investigations into the mechanisms that baculoviruses use to persist in natural insect populations.

 

Host cell interactions

Viruses are known to utilise the host cell cytoskeleton for trafficking within an infected cell, and both microtubules and actin filaments have been implicated in the replication cycle of baculoviruses. We have shown that the P10 protein of AcMNPV is closely associated with a dramatic rearrangement of microtubules in infected cells, and we propose the interaction plays a significant role in the late stages of virus occlusion. Furthermore, our studies have suggested a role for the actin microfilament network during the trafficking of virus glycoproteins to the plasma membrane. Disruption of these filaments severely curtailed the production or egress of intracellular budded virus, although nucleocapsid assembly and occlusion appeared independent of actin filament formation.
 
Current work in the group includes studying the interactions between baculovirus and the insect cell. We are utilising fluorescent protein tagged virus proteins and a range of organelle markers for live cell imaging, and higher resolution electron microscopy to uncover how the host cell changes during viral infection.
 
Baculovirus trafficking

We are studying the mechanisms of baculovirus movement in cells and in particular, the role of the cytoskeleton in the anterograde transport of virus from the nucleus to cell surface and in the detailed analysis of virus entry and exit from the nucleus. Recent work in the group has shown that AcMNPV proteins associate with kinesin-1 in insect cells, and that it is key in trafficking of virus from nucleus to plasma membrane prior to budding (Danquah et al. 2012). We continue to investigate anterograde virus trafficking using a range of microscopical approaches.

Understanding the P10 enigma

Our interest in the interaction of viruses with the host cell cytoskeleton has led us to investigate the role of the baculovirus P10 protein, a small protein known to interact with the cellular microtubule network. The gene encoding the P10 protein is evolutionarily conserved within all Alphabaculoviruses (nucleopolyhedroviruses of lepidopteran hosts), however, after over 3 decades of research its role in the baculovirus life cycle remains a mystery. P10 is non-essential for virus replication and spread, yet it is highly expressed very late in infection. We have recently demonstrated that P10 directly interacts with microtubules and can cause them to aggregate into thick bundles (Carpentier et al., 2008). P10 also self aggregates forming extensive perinuclear cytoskeletal like structures.
 
We have shown that upon their release from cells P10 strongly associates with polyhedral occlusion bodies. P10 may have a dual role; remodelling the microtubule cytoskeleton to benefit the virus replication cycle, and associating with occlusion bodies to enhance host to host transmission. These aspects of P10 are currently being investigated as is its potential as an enhancer of baculovirus-mediated transduction (due to its microtubule remodelling properties) during gene therapy. This research has also led us to become interested in looking at the mechanisms of virus occlusion and polyhedral occlusion body formation.
 
Development of novel baculovirus expression vectors
A number of approaches are being taken to improve the efficacy of baculoviruses as both vectors for the production of membrane targeted and secreted proteins, and as vectors for the control of insect pests. We are investigating ways of increasing the yield of recombinant protein production in stable insect cell lines and in novel vectors for the transduction of mammalian cells.
 

Baculoviruses as vectors for gene therapy

We are also working on the development of baculovirus vectors for gene expression in mammalian cells to ameliorate the effects of ischaemia reperfusion injury following transplantation.

People

  • Professor Linda A King
  • Dr Sarah Irons, Research Fellow in Cell Biology and Virology
  • Dr Caroline Griffiths, Senior Lecturer
  • Professor Robert D Possee, Visiting Professor
  • Professor Rosie Hails, Visiting Professor
  • Dr Adam Chambers, Visiting Researcher
  • Dr Richard Hitchman, Visiting Researcher
  • Farheen Raza, PhD student
  • Elisabetta Locanto, PhD student
  • Leo Graves, PhD student
  • Mine Aksular, PhD student

Funding

  • University
  • Industry
  • BBSRC studentships with The Pirbright Institute

Collaborations

  • Professor Chris Hawes (Oxford Brookes University)
  • Dr Casper Breuker (Oxford Brookes University)
  • Professor Isabel Bermudez (Oxford Brookes University)
  • Professor Robert Possee and Professor Rosie Hails (NERC Centre for Ecology and Hydrology, Oxford) 
  • Dr Ray Owens (Oxford Protein Production Facility, Research Complex at Harwell)
  • Professor George Rohrmann (Oregon State University, USA)
  • Professor Geoff Pilkington (University of Portsmouth)
  • Dr Stan Botchway (Central Laser Facility, Rutherford Appleton Laboratory)
  • Dr Javier Castillo-Olivares (The Pirbright Institute)
  • Dr David Staunton (Department of Biochemistry, University of Oxford)

Publications

Insect Virus Research Group Publications

CONTACT US

Professor Linda King
laking@brookes.ac.uk
+44 (0) 1865 483241 or
+44 (0) 1865 483600

Dr Caroline Griffiths
cgriffiths@brookes.ac.uk
+44 (0) 1865 484160

Insect virology microscopy

Insect virology microscopy