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Influenza virus-model membrane interaction. A morphological approach using modern cryotechniques
K N Burger1, G Knoll, A J Verkleij
1Institute of Molecular Biology and Medical Biotechnology, University of Utrecht, The Netherlands.
Abstract:
The membrane fusion activity of influenza virus was characterized morphologically using a model system composed of a highly purified influenza B virus suspension and ganglioside-containing zwitterionic liposomes. Electron microscopical analysis was performed after a combination of fast-freezing with either freeze-fracture or freeze-substitution-thin sectioning, ensuring maximal time resolution and avoiding preparation artifacts. In a parallel fluorescence 'lipid mixing' fusion assay, influenza virus-membrane fusion was characterized biochemically. Biochemical and morphological data are in full agreement, indicating negligible membrane fusion activity at neutral pH and high fusion activity at low pH. The freeze-fracture morphology strongly suggests a local point contact between viral and liposomal membrane at neutral pH, and a local point fusion mechanism for influenza virus-membrane fusion upon lowering of the pH. Fusion is followed by lipid mixing, lateral diffusion of viral spike proteins and exposure of viral contents at the inner liposomal surface.
Insights
Influenza B virus membrane fusion is minimal at neutral pH but highly active at low pH. Morphological and biochemical studies reveal a local point fusion mechanism, followed by lipid mixing and viral content release.
Area of Science:
- Virology
- Membrane Biology
- Biophysics
Background:
- Influenza virus entry into host cells is a critical step mediated by membrane fusion.
- Understanding the precise mechanism of viral membrane fusion is essential for developing antiviral strategies.
Purpose of the Study:
- To morphologically and biochemically characterize the membrane fusion activity of influenza B virus.
- To elucidate the mechanism of influenza virus-induced membrane fusion using a model system.
Main Methods:
- Utilized electron microscopy (freeze-fracture, freeze-substitution-thin sectioning) for high-resolution morphological analysis.
- Employed a fluorescence-based lipid mixing assay for biochemical characterization of fusion.
- Used a model system of purified influenza B virus and ganglioside-containing liposomes.
Main Results:
- Demonstrated negligible membrane fusion activity at neutral pH and significant fusion activity at low pH.
- Morphological data indicated a local point contact at neutral pH and a local point fusion mechanism at low pH.
- Biochemical and morphological findings were consistent, showing fusion followed by lipid mixing and viral content exposure.
Conclusions:
- Influenza B virus membrane fusion is pH-dependent, requiring acidic conditions for efficient activity.
- The fusion process initiates via a local point mechanism, leading to subsequent lipid mixing and viral content release into the liposome.