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Probing the spatiotemporal patterns of HBV multiplication reveals novel features of its subcellular processes

Through evolution, Hepatitis B Virus (HBV) developed highly intricate mechanisms exploiting host resources for its multiplication within a constrained genetic coding capacity. Yet a clear picture of viral hitchhiking of cellular processes with spatial resolution is still largely unsolved. Here, by l...

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Detalles Bibliográficos
Autores principales: Yue, Lei, Li, Chang, Xu, Mingzhu, Wu, Min, Ding, Jiahui, Liu, Jiangxia, Zhang, Xiaonan, Yuan, Zhenghong
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Public Library of Science 2021
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8376071/
https://www.ncbi.nlm.nih.gov/pubmed/34370796
http://dx.doi.org/10.1371/journal.ppat.1009838
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author Yue, Lei
Li, Chang
Xu, Mingzhu
Wu, Min
Ding, Jiahui
Liu, Jiangxia
Zhang, Xiaonan
Yuan, Zhenghong
author_facet Yue, Lei
Li, Chang
Xu, Mingzhu
Wu, Min
Ding, Jiahui
Liu, Jiangxia
Zhang, Xiaonan
Yuan, Zhenghong
author_sort Yue, Lei
collection PubMed
description Through evolution, Hepatitis B Virus (HBV) developed highly intricate mechanisms exploiting host resources for its multiplication within a constrained genetic coding capacity. Yet a clear picture of viral hitchhiking of cellular processes with spatial resolution is still largely unsolved. Here, by leveraging bDNA-based fluorescence in situ hybridization (FISH) combined with immunofluorescence, we developed a microscopic approach for multiplex detection of viral nucleic acids and proteins, which enabled us to probe some of the key aspects of HBV life cycle. We confirmed the slow kinetics and revealed the high variability of viral replication at single-cell level. We directly visualized HBV minichromosome in contact with acetylated histone 3 and RNA polymerase II and observed HBV-induced degradation of Smc5/6 complex only in primary hepatocytes. We quantified the frequency of HBV pregenomic RNAs occupied by translating ribosome or capsids. Statistics at molecular level suggested a rapid translation phase followed by a slow encapsidation and maturation phase. Finally, the roles of microtubules (MTs) on nucleocapsid assembly and virion morphogenesis were analyzed. Disruption of MTs resulted in the perinuclear retention of nucleocapsid. Meanwhile, large multivesicular body (MVB) formation was significantly disturbed as evidenced by the increase in number and decrease in volume of CD63(+) vesicles, thus inhibiting mature virion secretion. In conclusion, these data provided spatially resolved molecular snapshots in the context of specific subcellular activities. The heterogeneity observed at single-cell level afforded valuable molecular insights which are otherwise unavailable from bulk measurements.
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spelling pubmed-83760712021-08-20 Probing the spatiotemporal patterns of HBV multiplication reveals novel features of its subcellular processes Yue, Lei Li, Chang Xu, Mingzhu Wu, Min Ding, Jiahui Liu, Jiangxia Zhang, Xiaonan Yuan, Zhenghong PLoS Pathog Research Article Through evolution, Hepatitis B Virus (HBV) developed highly intricate mechanisms exploiting host resources for its multiplication within a constrained genetic coding capacity. Yet a clear picture of viral hitchhiking of cellular processes with spatial resolution is still largely unsolved. Here, by leveraging bDNA-based fluorescence in situ hybridization (FISH) combined with immunofluorescence, we developed a microscopic approach for multiplex detection of viral nucleic acids and proteins, which enabled us to probe some of the key aspects of HBV life cycle. We confirmed the slow kinetics and revealed the high variability of viral replication at single-cell level. We directly visualized HBV minichromosome in contact with acetylated histone 3 and RNA polymerase II and observed HBV-induced degradation of Smc5/6 complex only in primary hepatocytes. We quantified the frequency of HBV pregenomic RNAs occupied by translating ribosome or capsids. Statistics at molecular level suggested a rapid translation phase followed by a slow encapsidation and maturation phase. Finally, the roles of microtubules (MTs) on nucleocapsid assembly and virion morphogenesis were analyzed. Disruption of MTs resulted in the perinuclear retention of nucleocapsid. Meanwhile, large multivesicular body (MVB) formation was significantly disturbed as evidenced by the increase in number and decrease in volume of CD63(+) vesicles, thus inhibiting mature virion secretion. In conclusion, these data provided spatially resolved molecular snapshots in the context of specific subcellular activities. The heterogeneity observed at single-cell level afforded valuable molecular insights which are otherwise unavailable from bulk measurements. Public Library of Science 2021-08-09 /pmc/articles/PMC8376071/ /pubmed/34370796 http://dx.doi.org/10.1371/journal.ppat.1009838 Text en © 2021 Yue et al https://creativecommons.org/licenses/by/4.0/This is an open access article distributed under the terms of the Creative Commons Attribution License (https://creativecommons.org/licenses/by/4.0/) , which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
spellingShingle Research Article
Yue, Lei
Li, Chang
Xu, Mingzhu
Wu, Min
Ding, Jiahui
Liu, Jiangxia
Zhang, Xiaonan
Yuan, Zhenghong
Probing the spatiotemporal patterns of HBV multiplication reveals novel features of its subcellular processes
title Probing the spatiotemporal patterns of HBV multiplication reveals novel features of its subcellular processes
title_full Probing the spatiotemporal patterns of HBV multiplication reveals novel features of its subcellular processes
title_fullStr Probing the spatiotemporal patterns of HBV multiplication reveals novel features of its subcellular processes
title_full_unstemmed Probing the spatiotemporal patterns of HBV multiplication reveals novel features of its subcellular processes
title_short Probing the spatiotemporal patterns of HBV multiplication reveals novel features of its subcellular processes
title_sort probing the spatiotemporal patterns of hbv multiplication reveals novel features of its subcellular processes
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8376071/
https://www.ncbi.nlm.nih.gov/pubmed/34370796
http://dx.doi.org/10.1371/journal.ppat.1009838
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