Parvovirus B19 Uncoating Occurs in the Cytoplasm without Capsid Disassembly and It Is Facilitated by Depletion of Capsid-Associated Divalent Cations
Abstract
:1. Introduction
2. Materials and Methods
2.1. Cells and Viruses
2.2. Antibodies and Chemicals
2.3. Quantitative PCR
2.4. Iodixanol Density Gradient Ultracentrifugation
2.5. Transmission Electron Microscopy
2.6. Nuclease Assay
2.7. Isolation of Cytoplasmic and Nuclear Fractions from Infected Cells
2.8. Immunoprecipitation
2.9. Infectivity Assay
2.10. Complementary-Strand Synthesis
3. Results
3.1. Depletion of Capsid-Associated Divalent Cations Destabilizes the B19V Capsid
3.2. Depletion of Divalent Cations Facilitates B19V Uncoating without Capsid Disassembly
3.3. A Proportion of Incoming Capsids Uncoat in the Cytoplasm without Capsid Disassembly
3.4. Uncoating Occurs after Endosomal Escape
3.5. Incoming Capsids Become Reactive to Phosphospecific Antibodies Following Endosomal Escape
3.6. Capsids Phosphorylated and with Accessible Genomes Accumulated Progressively in the Nuclear Fraction
3.7. Nuclear Targeting of B19V Is Mediated by the Microtubule-Dependent, Minus-End-Directed Motor Dynein
3.8. Capsids Immunoprecipitated from Cytoplasmic and from Nuclear Fractions Support Complementary-Strand DNA Synthesis
4. Discussion
Author Contributions
Funding
Conflicts of Interest
References
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Caliaro, O.; Marti, A.; Ruprecht, N.; Leisi, R.; Subramanian, S.; Hafenstein, S.; Ros, C. Parvovirus B19 Uncoating Occurs in the Cytoplasm without Capsid Disassembly and It Is Facilitated by Depletion of Capsid-Associated Divalent Cations. Viruses 2019, 11, 430. https://doi.org/10.3390/v11050430
Caliaro O, Marti A, Ruprecht N, Leisi R, Subramanian S, Hafenstein S, Ros C. Parvovirus B19 Uncoating Occurs in the Cytoplasm without Capsid Disassembly and It Is Facilitated by Depletion of Capsid-Associated Divalent Cations. Viruses. 2019; 11(5):430. https://doi.org/10.3390/v11050430
Chicago/Turabian StyleCaliaro, Oliver, Andrea Marti, Nico Ruprecht, Remo Leisi, Suriyasri Subramanian, Susan Hafenstein, and Carlos Ros. 2019. "Parvovirus B19 Uncoating Occurs in the Cytoplasm without Capsid Disassembly and It Is Facilitated by Depletion of Capsid-Associated Divalent Cations" Viruses 11, no. 5: 430. https://doi.org/10.3390/v11050430
APA StyleCaliaro, O., Marti, A., Ruprecht, N., Leisi, R., Subramanian, S., Hafenstein, S., & Ros, C. (2019). Parvovirus B19 Uncoating Occurs in the Cytoplasm without Capsid Disassembly and It Is Facilitated by Depletion of Capsid-Associated Divalent Cations. Viruses, 11(5), 430. https://doi.org/10.3390/v11050430