The TRiC chaperonin controls reovirus replication through outer-capsid folding

Knowlton, Jonathan J.; Castro, Isabel Fernández de; Ashbrook, Alison W.; Gestaut, Daniel R.; Zamora, Paula F.; Bauer, Joshua A.; Forrest, J. Craig; Frydman, Judith; Risco, Cristina; Dermody, Terence S.

doi:10.1038/s41564-018-0122-x

Cited by 47 publications

(51 citation statements)

References 59 publications

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“…To rule out the possibility that the lower level of particle-associated 1 on purified particles is a consequence of 1 ejection due to the method of viral purification, we sought to measure the infectivity of unpurified, released viral progeny. Toward this goal, we used an assay recently described to identify host factors required for assembly and release of infectious progeny viruses (63). L929 cells were infected with equivalent attachment units of T3D F or T3D F /T3D C S1 for 24 h. Consistent with our data shown in Fig.…”

Section: Figsupporting

confidence: 55%

Protein Mismatches Caused by Reassortment Influence Functions of the Reovirus Capsid

Thete

Danthi

2018

J Virol

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Following attachment to host receptors via σ1, reovirus particles are endocytosed and disassembled to generate infectious subvirion particles (ISVPs). ISVPs undergo conformational changes to form ISVP*, releasing σ1 and membrane-targeting peptides from the viral μ1 protein. ISVP* formation is required for delivery of the viral core into the cytoplasm for replication. We characterized the properties of T3D/T3DS1, an S1 gene monoreassortant between two laboratory isolates of prototype reovirus strain T3D: T3D and T3D T3D/T3DS1 is poorly infectious. This deficiency is a consequence of inefficient encapsidation of S1-encoded σ1 on T3D/T3DS1 virions. Additionally, compared to T3D, T3D/T3DS1 undergoes ISVP-to-ISVP* conversion more readily, revealing an unexpected role for σ1 in regulating ISVP* formation. The σ1 protein is held within turrets formed by the λ2 protein. To test if the altered properties of T3D/T3DS1 are due to a mismatch between σ1 and λ2 proteins from T3D and T3D, properties of T3D/T3DL2 and T3D/T3DS1L2, which express a T3D-derived λ2, were compared. The presence of T3D λ2 allowed more efficient σ1 incorporation, producing particles that exhibit T3D-like infectivity. Compared to T3D, T3D/T3DL2 prematurely converts to ISVP*, uncovering a role for λ2 in regulating ISVP* formation. Importantly, a virus with matching σ1 and λ2 displayed a more regulated conversion to ISVP* than either T3D/T3DS1 or T3D/T3DL2. In addition to identifying new regulators of ISVP* formation, our results highlight that protein mismatches produced by reassortment can alter virus assembly and thereby influence subsequent functions of the virus capsid. Cells coinfected with viruses that possess a multipartite or segmented genome reassort to produce progeny viruses that contain a combination of gene segments from each parent. Reassortment places new pairs of genes together, generating viruses in which mismatched proteins must function together. To test if such forced pairing of proteins that form the virus shell or capsid alters the function of the particle, we investigated properties of reovirus variants in which the σ1 attachment protein and the λ2 protein that anchors σ1 on the particle are mismatched. Our studies demonstrate that a σ1-λ2 mismatch produces particles with lower levels of encapsidated σ1, consequently decreasing virus attachment and infectivity. The mismatch between σ1 and λ2 also altered the capacity of the viral capsid to undergo conformational changes required for cell entry. These studies reveal new functions of reovirus capsid proteins and illuminate both predictable and novel implications of reassortment.

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Section: Figsupporting

confidence: 55%

Protein Mismatches Caused by Reassortment Influence Functions of the Reovirus Capsid

Thete

Danthi

2018

J Virol

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“…A complex of these proteins and NS might be responsible for binding viral RNAs. It is also possible that NS binds cellular RNAs to promote the translation of cellular proteins within these structures (47), as several host proteins are found within inclusions, for example, Hsc70 (60) and the TRiC chaperonin (61).…”

Section: Discussionmentioning

confidence: 99%

Reovirus Nonstructural Protein σNS Acts as an RNA Stability Factor Promoting Viral Genome Replication

Zamora

Knowlton

et al. 2018

J Virol

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Viral nonstructural proteins, which are not packaged into virions, are essential for the replication of most viruses. Reovirus, a nonenveloped, double-stranded RNA (dsRNA) virus, encodes three nonstructural proteins that are required for viral replication and dissemination in the host. The reovirus nonstructural protein σNS is a single-stranded RNA (ssRNA)-binding protein that must be expressed in infected cells for production of viral progeny. However, the activities of σNS during individual steps of the reovirus replication cycle are poorly understood. We explored the function of σNS by disrupting its expression during infection using cells expressing a small interfering RNA (siRNA) targeting the σNS-encoding S3 gene and found that σNS is required for viral genome replication. Using complementary biochemical assays, we determined that σNS forms complexes with viral and nonviral RNAs. We also discovered, using and cell-based RNA degradation experiments, that σNS increases the RNA half-life. Cryo-electron microscopy revealed that σNS and ssRNAs organize into long, filamentous structures. Collectively, our findings indicate that σNS functions as an RNA-binding protein that increases the viral RNA half-life. These results suggest that σNS forms RNA-protein complexes in preparation for genome replication. Following infection, viruses synthesize nonstructural proteins that mediate viral replication and promote dissemination. Viruses from the family encode nonstructural proteins that are required for the formation of progeny viruses. Although nonstructural proteins of different viruses in the family diverge in primary sequence, they are functionally homologous and appear to facilitate conserved mechanisms of dsRNA virus replication. Using and cell culture approaches, we found that the mammalian reovirus nonstructural protein σNS binds and stabilizes viral RNA and is required for genome synthesis. This work contributes new knowledge about basic mechanisms of dsRNA virus replication and provides a foundation for future studies to determine how viruses in the family assort and replicate their genomes.

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“…In a later model, the RNA is proposed to be bound at the basic surface of a σ3 homodimer [193,194]. Recent data have actually confirmed the transition from σ3 homomultimer to structural σ3–μ1 heterohexamer by the action of cellular chaperonin [195].…”

Section: Protein Synthesis In Reovirus-infected Cellsmentioning

confidence: 99%

Synthesis and Translation of Viral mRNA in Reovirus-Infected Cells: Progress and Remaining Questions

Lemay

2018

Viruses

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At the end of my doctoral studies, in 1988, I published a review article on the major steps of transcription and translation during the mammalian reovirus multiplication cycle, a topic that still fascinates me 30 years later. It is in the nature of scientific research to generate further questioning as new knowledge emerges. Our understanding of these fascinating viruses thus remains incomplete but it seemed appropriate at this moment to look back and reflect on our progress and most important questions that still puzzle us. It is also essential of being careful about concepts that seem so well established, but could still be better validated using new approaches. I hope that the few reflections presented here will stimulate discussions and maybe attract new investigators into the field of reovirus research. Many other aspects of the viral multiplication cycle would merit our attention. However, I will essentially limit my discussion to these central aspects of the viral cycle that are transcription of viral genes and their phenotypic expression through the host cell translational machinery. The objective here is not to review every aspect but to put more emphasis on important progress and challenges in the field.

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The TRiC chaperonin controls reovirus replication through outer-capsid folding

Cited by 47 publications

References 59 publications

Protein Mismatches Caused by Reassortment Influence Functions of the Reovirus Capsid

Protein Mismatches Caused by Reassortment Influence Functions of the Reovirus Capsid

Reovirus Nonstructural Protein σNS Acts as an RNA Stability Factor Promoting Viral Genome Replication

Synthesis and Translation of Viral mRNA in Reovirus-Infected Cells: Progress and Remaining Questions

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