Origins of Enterovirus Replication Organelles Established by Whole-Cell Electron Microscopy
ABSTRACT Enterovirus genome replication occurs at virus-induced structures derived from cellular membranes and lipids. However, the origin of these replication organelles (ROs) remains uncertain. Ultrastructural evidence of the membrane donor is lacking, suggesting that the sites of its transition i...
| Main Authors: | , , , , , , , , |
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| Format: | Article |
| Language: | English |
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American Society for Microbiology
2019-06-01
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| Series: | mBio |
| Subjects: | |
| Online Access: | https://journals.asm.org/doi/10.1128/mBio.00951-19 |
| _version_ | 1818424661896069120 |
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| author | Charlotte E. Melia Christopher J. Peddie Anja W. M. de Jong Eric J. Snijder Lucy M. Collinson Abraham J. Koster Hilde M. van der Schaar Frank J. M. van Kuppeveld Montserrat Bárcena |
| author_facet | Charlotte E. Melia Christopher J. Peddie Anja W. M. de Jong Eric J. Snijder Lucy M. Collinson Abraham J. Koster Hilde M. van der Schaar Frank J. M. van Kuppeveld Montserrat Bárcena |
| author_sort | Charlotte E. Melia |
| collection | DOAJ |
| description | ABSTRACT Enterovirus genome replication occurs at virus-induced structures derived from cellular membranes and lipids. However, the origin of these replication organelles (ROs) remains uncertain. Ultrastructural evidence of the membrane donor is lacking, suggesting that the sites of its transition into ROs are rare or fleeting. To overcome this challenge, we combined live-cell imaging and serial block-face scanning electron microscopy of whole cells to capture emerging enterovirus ROs. The first foci of fluorescently labeled viral protein correlated with ROs connected to the endoplasmic reticulum (ER) and preceded the appearance of ROs stemming from the trans-Golgi network. Whole-cell data sets further revealed striking contact regions between ROs and lipid droplets that may represent a route for lipid shuttling to facilitate RO proliferation and genome replication. Our data provide direct evidence that enteroviruses use ER and then Golgi membranes to initiate RO formation, demonstrating the remarkable flexibility with which enteroviruses usurp cellular organelles. IMPORTANCE Enteroviruses are causative agents of a range of human diseases. The replication of these viruses within cells relies on specialized membranous structures termed replication organelles (ROs) that form during infection but whose origin remains elusive. To capture the emergence of enterovirus ROs, we use correlative light and serial block-face scanning electron microscopy, a powerful method to pinpoint rare events in their whole-cell ultrastructural context. RO biogenesis was found to occur first at ER and then at Golgi membranes. Extensive contacts were found between early ROs and lipid droplets (LDs), which likely serve to provide LD-derived lipids required for replication. Together, these data establish the dual origin of enterovirus ROs and the chronology of their biogenesis at different supporting cellular membranes. |
| first_indexed | 2024-12-14T14:01:36Z |
| format | Article |
| id | doaj.art-060c6b968b3649048287ffe1bae24e3f |
| institution | Directory Open Access Journal |
| issn | 2150-7511 |
| language | English |
| last_indexed | 2024-12-14T14:01:36Z |
| publishDate | 2019-06-01 |
| publisher | American Society for Microbiology |
| record_format | Article |
| series | mBio |
| spelling | doaj.art-060c6b968b3649048287ffe1bae24e3f2022-12-21T22:58:42ZengAmerican Society for MicrobiologymBio2150-75112019-06-0110310.1128/mBio.00951-19Origins of Enterovirus Replication Organelles Established by Whole-Cell Electron MicroscopyCharlotte E. Melia0Christopher J. Peddie1Anja W. M. de Jong2Eric J. Snijder3Lucy M. Collinson4Abraham J. Koster5Hilde M. van der Schaar6Frank J. M. van Kuppeveld7Montserrat Bárcena8Section Electron Microscopy, Department of Cell and Chemical Biology, Leiden University Medical Center, Leiden, The NetherlandsElectron Microscopy STP, The Francis Crick Institute, London, United KingdomSection Electron Microscopy, Department of Cell and Chemical Biology, Leiden University Medical Center, Leiden, The NetherlandsMolecular Virology Laboratory, Department of Medical Microbiology, Center of Infectious Diseases, Leiden University Medical Center, Leiden, The NetherlandsElectron Microscopy STP, The Francis Crick Institute, London, United KingdomSection Electron Microscopy, Department of Cell and Chemical Biology, Leiden University Medical Center, Leiden, The NetherlandsVirology Division, Faculty of Veterinary Medicine, Department of Infectious Diseases & Immunology, Utrecht University, Utrecht, The NetherlandsVirology Division, Faculty of Veterinary Medicine, Department of Infectious Diseases & Immunology, Utrecht University, Utrecht, The NetherlandsSection Electron Microscopy, Department of Cell and Chemical Biology, Leiden University Medical Center, Leiden, The NetherlandsABSTRACT Enterovirus genome replication occurs at virus-induced structures derived from cellular membranes and lipids. However, the origin of these replication organelles (ROs) remains uncertain. Ultrastructural evidence of the membrane donor is lacking, suggesting that the sites of its transition into ROs are rare or fleeting. To overcome this challenge, we combined live-cell imaging and serial block-face scanning electron microscopy of whole cells to capture emerging enterovirus ROs. The first foci of fluorescently labeled viral protein correlated with ROs connected to the endoplasmic reticulum (ER) and preceded the appearance of ROs stemming from the trans-Golgi network. Whole-cell data sets further revealed striking contact regions between ROs and lipid droplets that may represent a route for lipid shuttling to facilitate RO proliferation and genome replication. Our data provide direct evidence that enteroviruses use ER and then Golgi membranes to initiate RO formation, demonstrating the remarkable flexibility with which enteroviruses usurp cellular organelles. IMPORTANCE Enteroviruses are causative agents of a range of human diseases. The replication of these viruses within cells relies on specialized membranous structures termed replication organelles (ROs) that form during infection but whose origin remains elusive. To capture the emergence of enterovirus ROs, we use correlative light and serial block-face scanning electron microscopy, a powerful method to pinpoint rare events in their whole-cell ultrastructural context. RO biogenesis was found to occur first at ER and then at Golgi membranes. Extensive contacts were found between early ROs and lipid droplets (LDs), which likely serve to provide LD-derived lipids required for replication. Together, these data establish the dual origin of enterovirus ROs and the chronology of their biogenesis at different supporting cellular membranes.https://journals.asm.org/doi/10.1128/mBio.00951-19replication organelle biogenesiscoxsackieviruspicornavirusserial block-face scanning electron microscopySBF-SEMcorrelative light and electron microscopy |
| spellingShingle | Charlotte E. Melia Christopher J. Peddie Anja W. M. de Jong Eric J. Snijder Lucy M. Collinson Abraham J. Koster Hilde M. van der Schaar Frank J. M. van Kuppeveld Montserrat Bárcena Origins of Enterovirus Replication Organelles Established by Whole-Cell Electron Microscopy mBio replication organelle biogenesis coxsackievirus picornavirus serial block-face scanning electron microscopy SBF-SEM correlative light and electron microscopy |
| title | Origins of Enterovirus Replication Organelles Established by Whole-Cell Electron Microscopy |
| title_full | Origins of Enterovirus Replication Organelles Established by Whole-Cell Electron Microscopy |
| title_fullStr | Origins of Enterovirus Replication Organelles Established by Whole-Cell Electron Microscopy |
| title_full_unstemmed | Origins of Enterovirus Replication Organelles Established by Whole-Cell Electron Microscopy |
| title_short | Origins of Enterovirus Replication Organelles Established by Whole-Cell Electron Microscopy |
| title_sort | origins of enterovirus replication organelles established by whole cell electron microscopy |
| topic | replication organelle biogenesis coxsackievirus picornavirus serial block-face scanning electron microscopy SBF-SEM correlative light and electron microscopy |
| url | https://journals.asm.org/doi/10.1128/mBio.00951-19 |
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