Viruses and vesicles – cryo-electron microscopy and tomography
3D reconstruction of HIV-1 virions using cryo-electron microscopy (Briggs et al. 2006)
Correlated fluorescence and electron microscopy can be used to locate an individual fluorescent virus particle at the surface of a cell (Kukulski et al. 2011)
A high-resolution cryo-EM structure of an immature retrovirus capsid protein dimer (grey), fitted with available X-ray structures (Bharat et al. 2012)
A comparison of the arrangement of a retrovirus capsid protein in the immature (top) and mature (bottom) virus particles, showing dramatic conformational change (Bharat et al. 2012)
The Briggs group uses cryo-electron microscopy techniques to explore the mechanisms of assembly and budding of enveloped viruses and coated vesicles.
Previous and current research
We aim to understand the structure and molecular assembly mechanisms of important pathogenic enveloped viruses (e.g. HIV and Influenza virus), and of cellular trafficking vesicles (e.g. clathrin, COPI or COPII coated vesicles). To do this we need to understand how the protein components of the virus or vesicle interact with one another, how the cargo of the virus or vesicle is collected, and how the proteins interact with the lipid bilayer to reshape it and form a free virus or vesicle. The level of understanding we aim to achieve could be imagined as a 3D, functionally annotated movie, with molecular resolution, of the assembly and budding process. We apply methods including cryo-electron microscopy and tomography, correlated light and electron microscopy, and computational image processing. The unique power of cryo-electron microscopy and tomography methods is that they can provide detailed structural information under close-to-native conditions, even within cells.
For many problems we take a step-by-step approach. Correlative fluorescence and electron microscopy methods can be used to locate and characterise features of interest. 3D reconstructions of these features can be obtained using electron tomography of the biological system in its native state. These reconstructions can be better interpreted by comparison with data collected from in vitro reconstituted systems. A detailed view is obtained by carrying out image processing using sub-tomogram averaging or single-particle reconstruction. As well as applying existing methods, we develop and apply new approaches, in particular correlative fluorescence and electron microscopy techniques, and image processing protocols for high-resolution sub tomogram averaging. Members of the group have varied and complementary skills, including biochemistry, cell biology, physics, engineering and computer sciences.
A particular emphasis of our research is the structure and life-cycle of asymmetric membrane viruses such as HIV. The structure and assembly of HIV particles offers insights into general features of membrane budding. Further details on our research into the structure and inhibition of HIV are available on our Molecular Medicine Partnership Unit webpage.
Future projects and goals
Our goal is to understand the interplay between protein complexes, membrane shape and virus/vesicle structure. What kind of protein-protein interactions can drive virus assembly while maintaining structural flexibility? How do structural switches allow viruses and vesicles that have completed the assembly pathway to start disassembling? How do proteins induce the distortion of cellular membranes into vesicles of different dimensions? What are the similarities and differences between the variety of cellular budding events? How do viruses hijack cellular systems for their own use? How does the curvature of a membrane influence its interaction with particular proteins? We are developing and applying novel microscopy and image processing approaches to address these questions.
3D reconstructions of COPI coated vesicles obtained using cryo-electron tomography and sub-tomogram averaging (Faini et al. 2012)