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Proteolytic activation of tick-borne encephalitis virus by furin
K Stadler1, S L Allison, J Schalich
1Institute of Virology, University of Vienna, Austria.
Abstract:
Flaviviruses are assembled intracellularly in an immature form containing heterodimers of two envelope proteins, E and prM. Shortly before the virion exits the cell, prM is cleaved by a cellular enzyme, and this processing step can be blocked by treatment with agents that raise the pH of exocytic compartments. We carried out in vivo and in vitro studies with tick-borne encephalitis (TBE) virus to investigate the possible role of furin in this process as well as the functional consequences of prM cleavage. We found that prM in immature virions can be correctly cleaved in vitro by recombinant bovine furin but that efficient cleavage occurs only after exposure of the virion to mildly acidic pH. The data suggest that exposure to an acidic environment induces an irreversible structural change that renders the cleavage site accessible to the enzyme. Cleavage by furin in vitro resulted in biological activation, as shown by a 100-fold increase in specific infectivity, the acquisition of membrane fusion and hemagglutination activity, and the ability of the envelope proteins to undergo low-pH-induced structural rearrangements characteristic of mature virions. In vivo, prM cleavage was blocked by a furin inhibitor, and infection of the furin-deficient cell line LoVo yielded only immature virions, suggesting that furin is essential for cleavage activation of flaviviruses.
Insights
Furin enzyme is essential for activating flaviviruses, like tick-borne encephalitis virus, by cleaving the prM protein. This cleavage, triggered by acidic environments, is crucial for viral infectivity and maturation.
Area of Science:
- Virology
- Molecular Biology
- Biochemistry
Background:
- Flaviviruses assemble as immature virions with envelope proteins E and prM.
- prM cleavage by a cellular enzyme is required before virion exit.
- This processing step is sensitive to pH changes in exocytic compartments.
Purpose of the Study:
- Investigate the role of furin in prM cleavage for tick-borne encephalitis virus (TBEV).
- Determine the functional consequences of prM cleavage on viral maturation and infectivity.
Main Methods:
- In vitro cleavage assays using recombinant bovine furin.
- In vivo studies using furin inhibitors and furin-deficient cell lines (LoVo).
- Assays to measure viral infectivity, membrane fusion, hemagglutination, and protein structural changes.
Main Results:
- Recombinant furin cleaved TBEV prM in vitro, but only after exposure to mildly acidic pH.
- Acidic pH induced an irreversible structural change, making the cleavage site accessible.
- In vitro cleavage by furin significantly increased viral infectivity (100-fold) and induced mature virion characteristics.
- Furin inhibition in vivo blocked prM cleavage, and furin-deficient cells produced only immature virions.
Conclusions:
- Furin is essential for the cleavage activation of flaviviruses.
- Mildly acidic environments are necessary to trigger structural changes for efficient furin-mediated prM cleavage.
- prM cleavage by furin is a critical step for flavivirus maturation, infectivity, and the acquisition of functional properties.