Kryo-elektronenmikroskopische Analyse der Struktur und Dynamik von Proteinkomplexen bei der zellulären Infektion für Herpes simplex
Zusammenfassung der Projektergebnisse
In the course of the project we applied cryo electron tomography (cryo-ET) – a technique providing unprecedented insights into the cellular architecture at the macromolecular level – and complementary techniques to elucidated structural changes of macromolecular complexes in the course of the herpesvirus-host interaction. Being pioneers in cellular electron cryo tomography of host-pathogen interactions we established a number of dedicated cellular and subcellular investigation systems enabling the analysis of the interactions at different levels of complexity and resolution. These range from rather complex and close to native native systems like the use of dissociated hippocampal neurons for studying intracellular transport, via synaptosomes for virus entry studies, to liposome entry experiments and more reduced systems like display of isolated membrane proteins on liposomes or on heterologous pseudotyped viruses. These systems adjusted by the group for structural analyses are clearly also of interest and use to other researchers beyond the hostpathogen field. Using Herpes simplex virus 1, the causative agent of cold sores, as a model, we studied the involvement and interaction of macromolecules from both, the virus and the host cell in the course of infection. In that, we focussed on crucial steps in the Herpesvirus’ cellular ‘life cycle’ including virus entry, intracellular transport and virus assembly. By providing native structural information and novel insights into the mechanism of e.g. membrane fusion, we not only contribute to basic research in a human infectious disease, but likewise contribute to understanding central cell biological processes like cellular membrane flow or neurotransmitter release. Integrating this information with results from other techniques enabled a functional characterisation of the underlying dynamic processes. The results published in leading journals have led to a number of follow up research based on both, the technological advances and the biological insights.
Projektbezogene Publikationen (Auswahl)
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(2006) Cryoelectron tomographic structure of an immunodeficiency virus envelope complex in situ. PLoS Pathog. 2: e83
Zanetti, G., Briggs, J.A., Grünewald, K., Sattentau, Q.J., Fuller, S.D.
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(2006) The mechanism of HIV-1 core assembly: insights from three-dimensional reconstructions of authentic virions. Structure 14: 15-20
Briggs, J.A.G., Grünewald, K., Glass, B., Förster, F, Kräusslich, H.G., Fuller, S.D.
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(2007) Simian Virus 40 depends on ER protein folding and quality control factors for entry into host cells. Cell 131: 516-529
Schelhaas, M., Pelkmans, L., Malmström, J., Aebersold, R., Haugstetter, J., Ellgaard, L., Grünewald, K., Helenius, A.
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(2008) Insights into Bunyavirus architecture from electron cryo-tomography of Uukuniemi virus. PNAS 105: 2375-2379
Överby, A., Pettersson, R., Grünewald, K., Huiskonen, J.T.
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(2008) Intermediates of membrane fusion in herpesvirus entry captured by cryo electron tomography. PNAS 105(30): 10559- 10564
Maurer, U.E., Sodeik, B., Grünewald, K.
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(2010) Electron cryotomography of Tula hantavirus suggests a unique assembly paradigm for enveloped viruses. J. Virol. 84: 4889-4897
Huiskonen, J.T., Hepojoki, J., Laurinmäki, P., Vaheri, A., Lankinen, H., Butcher, S.J., Grünewald, K.
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(2011) Cryo Electron Tomography of Herpes Simplex Virus during Axonal Transport and Secondary Envelopment in Primary Neurons. PLoS Pathog. 7: e1002406
Ibiricu, I., Huiskonen, J.T., Döhner, K., Bradke, F., Sodeik, B., Grünewald, K.
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(2012) Correlative VIS-fluorescence and soft X-ray cryo-microscopy/tomography of adherent cells. J. Struct. Biol. 177: 193-201
Hagen, C., Guttmann, P., Klupp, B., Werner, S., Rehbein, S., Mettenleiter, T.C., Schneider, G., Grünewald, K.
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(2013) Characterization of herpes simplex virus type 1 L-particle assembly and egress in hippocampal neurons by electron cryotomography. Cell Microbiol. 15(2): 285-291
Ibiricu, I., Maurer, U.E., Grünewald, K.
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(2013) The structure of herpesvirus fusion glycoprotein B - bilayer complex reveals the protein-membrane and lateral protein-protein interaction. Structure 21: 1396-1405
Maurer, U.E., Zeev-Ben-Mordehai, T., Pandurangan, A.P., Cairns, T.M., Hannah, B.P., Whitbeck, J.C., Eisenberg, R.J., Cohen, G.H., Topf, M., Huiskonen, J.T., Grünewald, K.
