The Highly Ordered Double-Stranded RNA Genome of Bluetongue Virus Revealed by Crystallography

Gouet, Patrice; Diprose, Jonathan M.; Grimes, Jonathan M.; Malby, Robyn L.; Burroughs, J. N.; Zientara, Stéphan; Stuart, David I.; Mertens, Peter P. C.

doi:10.1016/s0092-8674(00)80758-8

Cited by 178 publications

(137 citation statements)

References 39 publications

(6 reference statements)

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“…The enlarged volume under the 12 fivefold vertices is about 9.4 × 10 5 Å 3 in total and accounts for 1.4% of the total volume of the capsid chamber. Although the transcription mechanism of the Reoviridae family has not been fully elucidated, a generally accepted transcription model is that RdRp is anchored to the capsid shell while a template RNA slides through RdRp for mRNA transcription (4,26,27). The enlarged capsid chamber enables genomic RNA to have greater flexibility to slide within the densely packed capsid.…”

Section: Conformational Changes In Capsid Shell Protein Vp1 and Relatedmentioning

confidence: 99%

Cryo-EM structure of a transcribing cypovirus

Yang

Liu

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

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Double-stranded RNA viruses in the family Reoviridae are capable of transcribing and capping nascent mRNA within an icosahedral viral capsid that remains intact throughout repeated transcription cycles. However, how the highly coordinated mRNA transcription and capping process is facilitated by viral capsid proteins is still unknown. Cypovirus provides a good model system for studying the mRNA transcription and capping mechanism of viruses in the family Reoviridae . Here, we report a full backbone model of a transcribing cypovirus built from a near-atomic-resolution density map by cryoelectron microscopy. Compared with the structure of a nontranscribing cypovirus, the major capsid proteins of transcribing cypovirus undergo a series of conformational changes, giving rise to structural changes in the capsid shell: ( i ) an enlarged capsid chamber, which provides genomic RNA with more flexibility to move within the densely packed capsid, and ( ii ) a widened peripentonal channel in the capsid shell, which we confirmed to be a pathway for nascent mRNA. A rod-like structure attributable to a partially resolved nascent mRNA was observed in this channel. In addition, conformational change in the turret protein results in a relatively open turret at each fivefold axis. A GMP moiety, which is transferred to 5’-diphosphorylated mRNA during the mRNA capping reaction, was identified in the pocket-like guanylyltransferase domain of the turret protein.

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Section: Conformational Changes In Capsid Shell Protein Vp1 and Relatedmentioning

confidence: 99%

Cryo-EM structure of a transcribing cypovirus

Yang

Liu

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

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“…Nascent mRNA is capped before being released from the intact capsid. The mechanism by which this characteristic endogenous transcription takes place inside intact dsRNA viruses has been studied extensively (3)(4)(5)(6)(7), and it is generally believed that the RNA polymerase complex is structurally anchored to the core during transcription (7)(8)(9). The dsRNA template must be flexible enough to move freely within the densely packed core so that it can slide through the RNAdependent RNA polymerase (RDRP) and the capping enzyme complex to ensure efficient transcription (10).…”

Section: Cpvmentioning

confidence: 99%

“…Both x-ray crystallography and electron cryomicroscopy (cryo-EM) have been employed to study the three-dimensional structures of dsRNA viruses. In particular, the crystal structure of the animal reovirus core (15), which is structurally homologous to CPV, and that of the bluetongue virus sub-core (7,16) have provided detailed descriptions of the structural and functional organizations of shell proteins. Yet due to the unavailability of the crystal structures of empty capsids, the interactions of RNA/protein (both TEC and shell protein) cannot be discerned directly in either case.…”

Section: Cpvmentioning

confidence: 99%

“…Previously, crystallography studies on the cores of animal reovirus and bluetongue virus have resolved their structures to atomic resolution (7,15). However, due to the unavailability of the empty capsid structures at the same resolution, the genomic RNA densities and their interactions with viral proteins could not be unambiguously resolved.…”

Section: Three-dimensional Structural Comparisons Between Fullmentioning

confidence: 99%

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Structural Comparisons of Empty and Full Cytoplasmic Polyhedrosis Virus

Xia¹,

Jakana²,

Zhang³

et al. 2003

Journal of Biological Chemistry

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Viruses in the family Reoviridae are capable of transcription within the intact capsids. As the only singleshelled and thus the simplest member of the Reoviridae, cytoplasmic polyhedrosis virus (CPV) provides an attractive system for studying endogenous transcription. We report the structures of the full and empty CPV determined at 13-Å resolution by electron cryomicroscopy. The structure of the empty CPV reveals a density attributed to the transcription enzyme complex, which is attached to the internal surface of the capsid shell below each of the 12 turrets. The full capsid has an identical capsid shell but contains additional internal densities contributed by the genomic double-stranded (ds) RNA. The RNA densities proximal to the capsid shell are organized into layers with a dodecahedral appearance, suggesting a genome organization of dsRNA segments each having a cone shape spooling around a transcription enzyme complex. Our structures also suggest that the capsid shell serves as a scaffold for appropriate positioning of the RNA genome, whereas nascent mRNA release takes place through the constricted central channel of the turret. Based on these observations, a detailed moving template transcription mechanism is proposed that may provide insight into the well coordinated and highly efficient endogenous RNA transcription of dsRNA viruses.

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“…A consequence of this is that the strategy of packaging a number of molecules which substantially exceeds the total number of distinct segments to achieve at least one copy of each with high probability is not a possibility (Gouet et al, 1999). Under conditions of transmission at high m.o.i.…”

Section: Introductionmentioning

confidence: 99%

Inter-segment complementarity in orbiviruses: a driver for co-ordinated genome packaging in the Reoviridae?

Boyce

McCrae

Boyce

et al. 2016

Journal of General Virology

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The process by which eukaryotic viruses with segmented genomes select a complete set of genome segments for packaging into progeny virus particles is not understood. In this study a model based on the association of genome segments through specific RNA-RNA interactions driven by base pairing was formalized and tested in the Orbivirus genus of the Reoviridae family. A strategy combining screening of the genomic sequences for inter-segment complementarity with direct functional testing of inter-segment RNA-RNA interactions using reverse genetics is described in the type species of the Orbivirus genus, Bluetongue virus (BTV). Two examples, involving four of the ten BTV genomic segments, of specific intersegment interaction motifs whose maintenance is essential for the generation of infectious virus, were identified. Equivalent inter-segment complementarities were found between the identified regions of the orthologous genome segments of all orbiviruses, including phylogenetically distant species. Specific interaction of the participating RNA segments was confirmed in vitro using electrophoretic mobility shift assays, with the interactions inhibited using oligonucleotides complementary to the interaction motif of one of the interacting partners, and also through mutagenesis of the motifs. In each example, the base pairing rather than the absolute sequence was critical to the formation of a functional inter-segment interaction, with mutations only being tolerated in rescued virus if compensating changes were made in the interacting partner to restore uninterrupted base pairing. The absolute sequence of the complementarity motifs varied between species, indicating that this newly identified phenomenon may contribute to the observed lack of reassortment between Orbivirus species.

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The Highly Ordered Double-Stranded RNA Genome of Bluetongue Virus Revealed by Crystallography

Cited by 178 publications

References 39 publications

Cryo-EM structure of a transcribing cypovirus

Cryo-EM structure of a transcribing cypovirus

Structural Comparisons of Empty and Full Cytoplasmic Polyhedrosis Virus

Inter-segment complementarity in orbiviruses: a driver for co-ordinated genome packaging in the Reoviridae?

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