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Multiple virulence determinants of foot-and-mouth disease virus in cell culture
E Baranowski1, N Sevilla, N Verdaguer
1Centro de Biología Molecular "Severo Ochoa", Universidad Autónoma de Madrid, 28049 Madrid, Spain.
Abstract:
Hypervirulent variants of foot-and-mouth disease virus (FMDV) of serotype C arise upon serial cytolytic or persistent infections in cell culture. A specific mutation in the internal ribosome entry site of persistent FMDV was previously associated with enhanced translation initiation activity that could contribute to the hypervirulent phenotype for BHK-21 cells. Here we report that several hypervirulent FMDV variants arising upon serial cytolytic passage show an invariant internal ribosome entry site but have a number of mutations affecting structural and nonstructural viral proteins. The construction of chimeric type O-type C infectious transcripts has allowed the mapping of a major determinant of hypervirulence to the viral capsid. Tissue culture-adapted FMDV displayed enhanced affinity for heparin, but binding to cell surface heparan sulfate moieties was not required for expression of the hypervirulent phenotype in Chinese hamster ovary (CHO) cells. Virulence was identical or even higher for glycosaminoglycan-deficient CHO cells than for wild-type CHO cells. FMDV variants with decreased affinity for heparin were selected from a high-binding parental population and analyzed. Substitutions associated with decreased heparin binding were located at positions 173 of capsid protein VP3 and 144 of capsid protein VP1. These substitutions had a moderate effect on virulence for BHK-21 cells but completely abrogated infection of CHO cells. The comparative results with several FMDV isolates show that (i) increased affinity for heparin and alterations in cell tropism may be mediated by a number of independent sites on the viral capsid and (ii) the same capsid modifications may have different effects on different cell types.
Insights
Hypervirulent foot-and-mouth disease virus (FMDV) variants emerge through mutations in viral capsid proteins, not just the internal ribosome entry site. These capsid changes influence heparin binding and alter cell tropism, impacting virulence differently across cell types.
Area of Science:
- Virology
- Molecular Biology
- Pathogenesis
Background:
- Hypervirulent variants of foot-and-mouth disease virus (FMDV) can emerge during cell culture adaptation.
- Previous studies linked mutations in the internal ribosome entry site (IRES) to enhanced translation and hypervirulence in specific cell lines (BHK-21).
Purpose of the Study:
- To investigate the genetic determinants of hypervirulence in FMDV arising from serial cytolytic passage.
- To identify viral components responsible for altered cell tropism and heparin binding in hypervirulent FMDV.
Main Methods:
- Construction of chimeric FMDV (type O-type C) infectious transcripts to map hypervirulence determinants.
- Analysis of FMDV variants with altered heparin-binding affinity.
- Assessment of viral infectivity and virulence in different cell lines (BHK-21, Chinese hamster ovary (CHO)) with varying glycosaminoglycan expression.
Main Results:
- Hypervirulent FMDV variants from cytolytic passage showed capsid protein mutations, independent of IRES changes.
- A major determinant of hypervirulence was mapped to the viral capsid.
- FMDV variants exhibited enhanced heparin affinity, but heparin binding was not essential for hypervirulence in CHO cells.
- Specific substitutions in VP3 (position 173) and VP1 (position 144) reduced heparin binding and abrogated CHO cell infection, while only moderately affecting BHK-21 cell virulence.
Conclusions:
- Hypervirulence and altered cell tropism in FMDV can be mediated by multiple, independent sites on the viral capsid.
- Capsid modifications can have differential effects on FMDV virulence and cell tropism across distinct cell types.