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Autographa californica M nuclear polyhedrosis virus: microtubules and replication
1Department of Entomology, University of California, Berkeley 94720.
Abstract:
Progressive reorganization and depolymerization of microtubules corresponded with virus-induced rounding of Autographa californica M nuclear polyhedrosis virus (AcMNPV)-infected Spodoptera frugiperda IPLB-Sf-21 cells, suggesting that microtubules were instrumental in maintaining the normal shape of these cells. Depolymerization of all cortical and most of the paranuclear microtubules with colchicine also resulted in cell rounding, confirming this hypothesis. Studies with aphidicolin and cycloheximide indicated the virus-induced effects on the microtubules were mediated by both early and late viral gene products. Microtubules in cells infected with a p10 deletion mutant depolymerized microtubules in a manner similar to those in wild-type virus-infected cells, indicating p10 was not responsible for virus-induced changes in the microtubules. Nevertheless, evidence for the association of p10 and microtubules was obtained by fluorescence microscopy and immunoelectron microscopy. Colchicine depolymerization of microtubules before and throughout infection did not interfere with virus replication, but treatment of cells with taxol, a microtubule-stabilizing agent, both delayed and depressed virus replication. The taxol-induced effect was relieved by the addition of colchicine. These results suggested that AcMNPV-induced depolymerization of microtubules may be a necessary event in, rather than a tangential effect of, virus replication. Attempts to monitor the effects of virus infection on intermediate filaments were unsuccessful due to the lack of cross-reactivity between antibodies to intermediate filament proteins and IPLB-Sf-21 cells, indicating these proteins are not highly conserved in lepidopteran insect cells.
Insights
Autographa californica M nuclear polyhedrosis virus (AcMNPV) infection causes microtubule depolymerization, essential for viral replication. Disrupting microtubules with colchicine did not affect replication, but stabilizing them with taxol inhibited it.
Area of Science:
- Cell biology
- Virology
- Insect molecular biology
Background:
- Microtubules maintain cell shape and are crucial cellular components.
- Autographa californica M nuclear polyhedrosis virus (AcMNPV) infects Spodoptera frugiperda cells.
- The role of cellular structures in AcMNPV replication is not fully understood.
Purpose of the Study:
- To investigate the role of microtubules in AcMNPV infection.
- To determine the viral mechanisms responsible for microtubule alterations.
- To assess the impact of microtubule dynamics on viral replication.
Main Methods:
- Cell rounding assays in AcMNPV-infected Spodoptera frugiperda cells.
- Treatment with microtubule-disrupting (colchicine) and stabilizing (taxol) agents.
- Use of aphidicolin and cycloheximide to study viral gene products.
- Infection with a p10 deletion mutant virus.
- Fluorescence and immunoelectron microscopy to examine protein-microtubule interactions.
Main Results:
- AcMNPV infection induced progressive microtubule reorganization and depolymerization, correlating with cell rounding.
- Colchicine-induced microtubule depolymerization also caused cell rounding, confirming microtubules' role in cell shape.
- Both early and late viral gene products mediated the virus-induced microtubule effects.
- A p10 deletion mutant affected microtubules similarly to wild-type virus, indicating p10 is not essential for these changes.
- Despite this, p10 showed association with microtubules.
- Colchicine treatment did not impede viral replication.
- Taxol treatment delayed and reduced viral replication, an effect reversed by colchicine.
- Intermediate filament studies were inconclusive due to antibody cross-reactivity issues.
Conclusions:
- Microtubule depolymerization is a necessary event for AcMNPV replication, not merely a side effect.
- Viral gene products, excluding p10, are responsible for mediating microtubule alterations.
- Microtubule dynamics play a critical role in the AcMNPV life cycle.