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Interaction of Theiler's virus with intermediate filaments of infected cells
P Nédellec1, P Vicart, C Laurent-Winter
1Unité des Virus Lents (ERS 572 CNRS), Département SIDA & Rétrovirus, Institut Pasteur, 75724 Paris Cedex 15, France.
Abstract:
Theiler's murine encephalomyelitis virus is a neurotropic murine picornavirus which replicates permissively and causes a cytopathic effect in the BHK-21 cell line. We examined the interactions between the GDVII and DA strains of Theiler's virus and BHK-21 host cell proteins in a virus overlay assay. We observed binding of the virions to two proteins of approximately 60 kDa. These proteins were microsequenced and identified as desmin and vimentin, two main components of the intermediate filament network. The association between desmin or vimentin and virions was demonstrated by immunoprecipitation. Anti-desmin and anti-vimentin monoclonal antibodies precipitated GDVII or DA virions from extracts of infected BHK-21 cells. The intracellular distributions of virions and of the desmin and vimentin intermediate filaments of BHK-21 cells were investigated by two-color immunofluorescence confocal microscopy. Following infection, the intermediate filament network was rearranged into a shell-like structure which surrounded a viral inclusion. Finally, close contact between GDVII virus particles and 10-nm intermediate filaments was observed by electron microscopy.
Insights
Theiler's virus (GDVII and DA strains) binds to desmin and vimentin, key intermediate filament proteins in BHK-21 cells. Viral infection rearranges these filaments, forming a shell around viral inclusions.
Area of Science:
- Virology
- Cell Biology
- Biochemistry
Background:
- Theiler's murine encephalomyelitis virus (TMEV) is a neurotropic picornavirus.
- BHK-21 cells are a common cell line used for viral replication studies.
- Intermediate filaments, including desmin and vimentin, are crucial cellular structural components.
Purpose of the Study:
- To investigate the interactions between TMEV strains (GDVII and DA) and host cell proteins in BHK-21 cells.
- To identify host cell proteins that bind to TMEV virions.
- To elucidate the effect of TMEV infection on the cellular intermediate filament network.
Main Methods:
- Virus overlay assay to detect virion-protein interactions.
- Microsequencing to identify bound proteins.
- Immunoprecipitation to confirm protein-virion association.
- Two-color immunofluorescence confocal microscopy to study intracellular distribution.
- Electron microscopy for high-resolution visualization.
Main Results:
- TMEV virions bound to two host proteins of approximately 60 kDa.
- These proteins were identified as desmin and vimentin, components of the intermediate filament network.
- Anti-desmin and anti-vimentin antibodies precipitated TMEV virions.
- TMEV infection induced rearrangement of the intermediate filament network into a shell-like structure surrounding viral inclusions.
- Electron microscopy revealed close contact between virus particles and intermediate filaments.
Conclusions:
- Desmin and vimentin are host cell proteins that interact with TMEV GDVII and DA strains.
- TMEV infection alters the organization of the intermediate filament network in BHK-21 cells.
- The observed rearrangement suggests a role for intermediate filaments in the TMEV replication cycle or cellular response to infection.