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African swine fever virus interaction with microtubules
1Pathology Department, Curry Cabral Hospital, Lisbon, Portugal.
Abstract:
The role of microtubules in intracellular transport of African swine fever virus (ASFV) and virus-induced inclusions was studied by immunofluorescence using anti-ASFV and anti-tubulin antibodies, by electron microscopy of infected Vero cells and by in vitro binding of virions to purified microtubules. MTC, a reversible colchicine analogue, was used to depolymerize microtubules. In cells treated with MTC multiple large inclusions containing ASFV antigens and particles were observed in the cytoplasm. Removal of the drug lead to migration and fusion of the inclusions at a perinuclear location. To study the effect of microtubule repolymerization on virus particle distribution, the particles were counted in thin sections of MTC treated cells and at different times after removal of the drug. In cells treated with MTC 6.8% and 3.6% of the virus particles were found respectively in the cytoplasm and at the cell membrane while 38% of the particles were located around the virosome. With reversal of the drug effect the number of virus particles around the virosomes progressively decreased to 10% at 2 h while the number of particles in the cytoplasm and at the cell membrane increased. At 2 h after removal of the drug 33.5% of the particles were found budding from the cell membrane. Virus particles were found closely associated with microtubules in cytoskeletons obtained by Triton X-100 extraction of taxol treated cells. The association of virus particles with microtubules was also observed in vitro using purified microtubules and virus particles.(ABSTRACT TRUNCATED AT 250 WORDS)
Insights
African swine fever virus (ASFV) intracellular transport relies on microtubules. Disrupting microtubules causes ASFV inclusions to accumulate, while their repolymerization facilitates virus transport and cell membrane budding.
Area of Science:
- Cell Biology
- Virology
Background:
- Microtubules play a crucial role in intracellular transport processes.
- Understanding viral transport mechanisms is key to controlling viral infections.
Purpose of the Study:
- To investigate the role of microtubules in the intracellular transport of African swine fever virus (ASFV).
- To examine the impact of microtubule depolymerization and repolymerization on ASFV particle distribution and cell membrane association.
Main Methods:
- Immunofluorescence microscopy using anti-ASFV and anti-tubulin antibodies.
- Electron microscopy of infected Vero cells.
- In vitro binding assays with purified microtubules and virions.
- Treatment with MTC (a colchicine analogue) to depolymerize microtubules.
Main Results:
- Depolymerization of microtubules with MTC led to the accumulation of ASFV inclusions in the cytoplasm.
- Reversal of MTC treatment resulted in the migration and perinuclear fusion of inclusions.
- Microtubule repolymerization correlated with decreased virus particles around virosomes and increased particles in the cytoplasm and at the cell membrane.
- ASFV particles were observed budding from the cell membrane after microtubule repolymerization.
- Direct association of ASFV particles with microtubules was confirmed both in vitro and in cell cytoskeletons.
Conclusions:
- Microtubules are essential for the intracellular trafficking of ASFV.
- The dynamic state of microtubules influences the distribution and egress of ASFV particles.
- Targeting microtubule-dependent transport could be a strategy for ASFV control.