Structure and arrangement of viral proteins in HIV, Marburg and Influenza viruses
Final Report Abstract
In this research programme, we used cryo-electron tomography and sub-tomogram averaging methods to study the structure of enveloped viruses including retroviruses and filoviruses. The irregular shapes of these viruses make them difficult to study by conventional structural biology methods. Our studies of retroviruses focused on HIV, with comparative studies on other retroviruses. HIV assembles in two stages, first forming an immature virus particle, and then undergoing proteolytic cleavage of viral proteins to change into a mature infectious virion. We described the arrangement of Gag (the major structural protein) in 3D within immature virus particles. This allowed us to describe how the proteins are positioned relative to one another to form a lattice within the immature virus. Surprisingly we found that the Gag lattice incorporates irregularly shaped defects in order to curve into a roughly spherical structure, and that the lattice contains one large gap at the point where the virus left the cell surface. Subsequent studies of assembled Gag protein from Mason-Pfizer monkey virus using a combination of cryo-electron microscopy and tomography allowed us to obtain the first highresolution structure of an immature retroviral Gag lattice, thereby defining protein interfaces involved in retrovirus assembly, and revealing the dramatic structural rearrangements that occur when the virus matures. Our studies of filoviruses encompassed both Marburg virus and Ebola virus. We used electron microscopy, cryo-electron microscopy, and cryo-electron tomography to image a range of different samples including virus-like particles assembled from purified proteins, virus particles, and infected cells. In combination these data allowed us to see the 3D structure of the rod-shaped genome-binding nucleocapsid, locate the different protein components within the nucleocapsid, and identify the minimal protein requirements for its assembly. We found that the nucleocapsid binds to the inside of the membrane of the infected cell in its final structural form and is wrapped by the membrane starting at one end. The filovirus nucleocapsid is similar to that of rhabdoviruses, but is wrapped from the opposite end, suggesting different ways of starting the wrapping process. Together these studies provide substantial insight into the 3D structure of these major human pathogens, illustrate how regular protein arrangements can provide a basis for virus particles that have irregular shape, and provide methods for the further study of the structure of irregular enveloped viruses.
Publications
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Structure and assembly of immature HIV. (2009) Proceedings of the National Academy of Sciences, 106, 11090-11095
Briggs, J.A.G., Riches, J.D., Glass, B., Bartonova, V., Zanetti, G. and Krausslich, H.G.
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Conserved and variable features of Gag structure and arrangement in immature retrovirus particles. (2010) Journal of Virology, 84, 11729-36
de Marco, A., Davey, N.E., Ulbrich, P., Phillips, J.M., Lux, V., Riches, J.D., Fuzik, T., Ruml, T., Krausslich, H.G., Vogt., V.M. and Briggs, J.A.G.
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Electron tomography reveals the steps in filovirus budding. (2010) PLoS Pathogens, 6, e1000875
Welsch, S., Kolesnikova, L., Kraehling, V, Riches, J.D., Becker, S. and Briggs, J.A.G.
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Structural Analysis of HIV-1 Maturation Using Cryo-Electron Tomography. (2010) PLoS Pathogens, 6, e1001215
de Marco, A., Müller, B., Glass, B., Riches, J.D., Kräusslich, H-G and Briggs, J.A.G.
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Cryo-Electron Tomography of Marburg Virus Particles and Their Morphogenesis within Infected Cells. (2011) PLoS Biology 9: e1001196
Bharat, T.A., Riches, J.D., Kolesnikova, L., Welsch, S., Krahling, V., Davey, N., Parsy, M.L., Becker, S. and Briggs, J.A.G.
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Structure of the immature retroviral capsid at 8Å resolution by cryo-electron microscopy. (2012) Nature, 487, 385-9
Bharat, T.A.M., Davey, N.E., Ulbrich, P., Riches, J.D., de Marco, A., Rumlova, M., Sachse, C., Ruml, T. and Briggs, J.A.G.
