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Monensin inhibits Semliki Forest virus penetration into culture cells
Abstract:
The carboxylic ionophores monensin and nigericin, at concentrations higher than 10 and 6 muM, respectively, prevent the penetration of the Semliki Forest virus (SFV) genome into the cytosol of baby hamster kidney (BHK-21) cells and thereby inhibit viral replication. In the absence of inhibitors, the entry of SFV is known to proceed by adsorptive endocytosis in coated vesicles, followed by acid-triggered membrane fusion in intracellular vacuoles or lysosomes. The results show that binding of the virus to the cell surface, adsorptive endocytosis, and intracellular transport of viruses to the lysosomes are only marginally affected by the ionophores. No direct virucidal effect is observed, nor is the membrane fusion activity of the virus at low pH directly affected. Sequential addition of monensin and ammonium chloride (a non-related lysosomotropic inhibitor of SFV entry) indicates that both inhibitors affect the same step in the entry pathway. On the basis of these data and the known effects of carboxylic ionophores and lysosomotropic weak bases on cellular pH gradients, we conclude that monensin inhibits penetration by increasing the pH in endocytic vacuoles and lysosomes above pH 6, which is the pH threshold for the viral membrane fusion activity.
Insights
Carboxylic ionophores like monensin inhibit Semliki Forest virus (SFV) replication by blocking genome entry into cells. These ionophores disrupt the pH gradient necessary for viral membrane fusion, a critical step in viral infection.
Area of Science:
- Virology
- Cell Biology
- Biochemistry
Background:
- Semliki Forest virus (SFV) enters host cells via adsorptive endocytosis.
- Viral entry involves acid-triggered membrane fusion within intracellular vacuoles.
- Understanding viral entry mechanisms is crucial for developing antiviral strategies.
Purpose of the Study:
- To investigate the effect of carboxylic ionophores monensin and nigericin on SFV entry and replication.
- To elucidate the specific step in the viral entry pathway inhibited by these ionophores.
Main Methods:
- Utilized baby hamster kidney (BHK-21) cells and Semliki Forest virus (SFV).
- Applied carboxylic ionophores monensin and nigericin at specific concentrations.
- Performed sequential addition experiments with monensin and ammonium chloride.
- Assessed viral genome penetration, replication, and membrane fusion activity.
Main Results:
- Monensin and nigericin inhibited SFV genome penetration and replication at concentrations above 10 muM and 6 muM, respectively.
- Ionophores had minimal impact on virus binding, endocytosis, and intracellular transport.
- No direct virucidal effect or inhibition of low pH-induced membrane fusion was observed.
- Sequential addition experiments suggested monensin and ammonium chloride target the same entry step.
Conclusions:
- Carboxylic ionophores inhibit SFV penetration by increasing the pH within endocytic vacuoles and lysosomes.
- This pH increase exceeds the threshold required for viral membrane fusion.
- Monensin acts as an inhibitor of viral entry by disrupting cellular pH gradients essential for fusion.