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Role of ATP in influenza virus budding
1Department of Microbiology, Immunology, and Molecular Genetics, UCLA School of Medicine, Los Angeles, California 90095-1747, USA.
Abstract:
Influenza viruses bud from the plasma membrane of virus-infected cells. Although budding is a critical step in virus replication, little is known about the requirements of the budding process. In this report, we have investigated the role of ATP in influenza virus budding by treating influenza virus infected Madin-Darby canine kidney (MDCK) cells with a number of metabolic inhibitors. When WSN virus-infected MDCK cells were exposed to antimycin A, carbonyl cyanide m-chlorophenylhydrazone, carbonyl cyanide p-trifluoromethoxy-phenylhydrazone, or oligomycin for a short time (15 min or 1 h) late in the infectious cycle, the rate of virus budding decreased. This inhibitory effect was reversible upon removal of the inhibitors. The role of ATP hydrolysis was analyzed by treating lysophosphatidylcholine (LPC)-permeabilized live filter-grown virus-infected MDCK cells with nonpermeable ATP analogues from the basal side and assaying virus budding from the apical side. In LPC-permeabilized cells, membrane-impermeable ATP analogues such as adenosine 5'-O-(3-thiotriphosphate) or 5'-adenylylimidodiphosphate caused reduction of virus budding which could be partially restored by adding excess ATP. These data demonstrated that ATP hydrolysis and not just ATP binding was required for virus budding. However, inhibitors of ion channel (ATPases) and protein ubiquitinylation, which also required the ATP as energy source, did not affect influenza virus budding, suggesting that neither ion channel nor protein ubiquitinylation activity was involved in influenza virus budding. On the other hand, treatment with dimethyl sulfoxide (DMSO), which decreases membrane viscosity, reduced the rate of virus budding, demonstrating that the physical state of membrane viscosity and membrane fluidity had an important effect on virus budding. Data presented in the report indicate that influenza virus budding is an active ATP-dependent process and suggest that reduced virus budding by ATP depletion and DMSO treatment may be partly due to decreased membrane viscosity.
Insights
Influenza virus budding requires ATP hydrolysis, not just binding. Metabolic inhibitors that deplete ATP reduce budding, which is reversible. Membrane viscosity also impacts this critical viral replication step.
Area of Science:
- Virology
- Cell Biology
- Biochemistry
Background:
- Influenza virus replication involves budding from host cell plasma membranes.
- The specific requirements and energy dependence of this budding process remain largely uncharacterized.
Purpose of the Study:
- To investigate the role of adenosine triphosphate (ATP) and its hydrolysis in influenza virus budding.
- To explore the influence of membrane biophysical properties on virus budding.
Main Methods:
- Treatment of infected Madin-Darby canine kidney (MDCK) cells with metabolic inhibitors affecting ATP production.
- Analysis of ATP hydrolysis requirement using lysophosphatidylcholine-permeabilized cells and non-permeable ATP analogues.
- Assessment of virus budding rates under varying membrane viscosity conditions using dimethyl sulfoxide (DMSO).
Main Results:
- Metabolic inhibitors targeting ATP production significantly reduced influenza virus budding in a reversible manner.
- ATP hydrolysis, but not ATP binding alone, was essential for efficient virus budding.
- Inhibitors of ion channels and protein ubiquitination did not affect budding, ruling out their direct involvement.
- Decreased membrane viscosity induced by DMSO treatment also inhibited virus budding.
Conclusions:
- Influenza virus budding is an active, ATP-dependent process relying on ATP hydrolysis.
- Membrane biophysical properties, specifically viscosity and fluidity, play a crucial role in regulating virus budding efficiency.