A 200 kV electron microscope for structural studies in the MRC Virology Unit

Lead Research Organisation: Medical Research Council
Department Name: Medical Research Council

Abstract

Viruses are among the smallest disease agents that infect humans, existing at the boundary between life and chemistry. The infectious particle, or ‘virion‘, consists of a genome (i.e. a collection of genes) and a shell that surrounds and protects the genome, and carries it from one host to the next. In many types of virus, the shell assembles from numerous copies of one or a few kinds of protein molecule. Many form in a symmetrical manner and are often helical or spherical in shape. Viruses do not reproduce on their own: instead, they invade the cells of their host and redirect the cell‘s resources to make new virus particles. To achieve this, an infecting virion must attach to the cell‘s surface and make its way into the cell‘s interior. The virus takes over control of the cell, switching off defence mechanisms and using cellular systems to read the virus genome and follow its instructions to make new virions. These new virus particles then exit the cell, and can infect another cell in the same host or a different host. As well as overcoming defences inside cells, viruses may also be capable of evading the host‘s immune system to some extent. In many cases, virus genes mutate rapidly, thus changing the appearance of the virion to the host‘s immune system. Distinct families of viruses may be very different in their structural and functional organisation.

To understand how viruses accomplish these complex processes, we need to study the structures of virions and viral components, both in isolation and when they interact with host cells. We need to visualise the structures in sufficient detail that we can start to form ideas about how the molecules making up the structures carry out their functions in the virus life cycle. One approach to analysing such structures is to use a very powerful electron microscope, and to process the images from the microscope using a computer. The high resolution views obtained in this way should then aid understanding ofthe virus?s molecular machinery work. With such knowledge, we can begin to devise ways of blocking or derailing the actions of the virus. For example, by understanding how viruses recognise and enter cells, we can envisage designing drugs to block entry. Combined with modern molecular biology techniques, this kind of research will provide a powerful weapon in our war against infectious diseases.

Technical Summary

The MRC Virology Unit represents a prominent part of MRC‘s investment in fundamental research into viral agents of human disease. The main viruses presently under study include cytomegalovirus, herpes simplex virus, hepatitis C virus and respiratory syncytial virus. In the area of structural research, Dr Frazer Rixon, programme leader in the Unit, has had a long-standing and highly productive collaboration with a group in Texas, on herpesvirus particle structure as studied by electron cryo-microscopy and computerised image reconstruction. The Unit has operated its own electron microscopy facility for many years, and this has been utilised in many aspects of its research. A project was started in 1996 to establish cryo-microscopy and image reconstruction capabilities in the Unit and to apply these techniques to investigating virions and virus components, with the aim of increasing understanding of virion structures and mechanics, and of interactions between viral and cellular elements. Now led by Dr David Bhella, this has achieved substantial successes, with results on herpesvirus particles, nucleocapsids of negative-stranded RNA viruses, and complexes between echovirus particles and receptors.

The further advance of structural studies in the Unit has become severely limited by the resolution of our cryo-capable microscope, which is a 120 kV instrument commissioned in 1994. We are therefore making this application to MRC to purchase a state-of-the-art, 200 kV transmission electron microscope. This will renew the Unit‘s electron microscopy facility for the foreseeable future, to a competent, modern level. It will support Rixon‘s and Bhella‘s investigations, and will be widely used by other Unit scientists. Specific plans concern maturation processes in herpesvirus capsids, fine structures of paramyxovirus nucleocapsids containing the virus transcriptional apparatus, high resolution analyses of enterovirus-receptor interactions, and investigation of hepatitis derived particles. The instrument will have a tomographic capability, which will be applied to investigation of single particles of complex virions and to exploring in three dimensions the interiors of virus-infected cells. We expect a considerable demand for time on this instrument from colleagues in biological and physical disciplines from Glasgow and beyond, and we shall facilitate such access. We shall commit a substantial sum from the Unit‘s allocated funds toward the purchase. In addition, the application has enthusiastic support from the Dean of the Faculty of Biomedical and Life Sciences, Glasgow University, and he has undertaken to contribute certain ancillary equipment and to prepare appropriate accommodation.

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