Structural features of the Bluetongue virus NS2 protein

Mumtsidu, Eleni; Makhov, Alexander M.; Roessle, Manfred; Bathke, Anja; Tucker, Paul A.

doi:10.1016/j.jsb.2007.07.013

Cited by 19 publications

(17 citation statements)

References 33 publications

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“…This represents a novel mechanism of modulating RNA chaperone activity, whereby the oligomeric state of the protein defines its unwinding efficiency. Similar to ARV σNS, mammalian reovirus σNS and bluetongue virus NS2 have also been reported to exist in a range of oligomeric states (63–66). It is possible that different σNS oligomers may play distinct roles during viral replication.…”

Section: Discussionmentioning

confidence: 99%

Stability of local secondary structure determines selectivity of viral RNA chaperones

Bravo

Borodavka

Barth

et al. 2018

Nucleic Acids Research

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To maintain genome integrity, segmented double-stranded RNA viruses of the Reoviridae family must accurately select and package a complete set of up to a dozen distinct genomic RNAs. It is thought that the high fidelity segmented genome assembly involves multiple sequence-specific RNA–RNA interactions between single-stranded RNA segment precursors. These are mediated by virus-encoded non-structural proteins with RNA chaperone-like activities, such as rotavirus (RV) NSP2 and avian reovirus σNS. Here, we compared the abilities of NSP2 and σNS to mediate sequence-specific interactions between RV genomic segment precursors. Despite their similar activities, NSP2 successfully promotes inter-segment association, while σNS fails to do so. To understand the mechanisms underlying such selectivity in promoting inter-molecular duplex formation, we compared RNA-binding and helix-unwinding activities of both proteins. We demonstrate that octameric NSP2 binds structured RNAs with high affinity, resulting in efficient intramolecular RNA helix disruption. Hexameric σNS oligomerizes into an octamer that binds two RNAs, yet it exhibits only limited RNA-unwinding activity compared to NSP2. Thus, the formation of intersegment RNA–RNA interactions is governed by both helix-unwinding capacity of the chaperones and stability of RNA structure. We propose that this protein-mediated RNA selection mechanism may underpin the high fidelity assembly of multi-segmented RNA genomes in Reoviridae.

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Section: Discussionmentioning

confidence: 99%

Stability of local secondary structure determines selectivity of viral RNA chaperones

Bravo

Borodavka

Barth

et al. 2018

Nucleic Acids Research

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show abstract

“…However, blocking of the C terminus with a V5 epitope tag precluded assembly into the larger aggregates characteristic of authentic VIBs, implying that the C terminus must be exposed to allow correct VIB formation. Previous structural studies suggesting a helical oligomeric structure for NS2 aggregates in which the C terminus is involved in the intermolecular interactions may help explain the necessity for the C terminus exposure, as well as the lack of aggregated VIB-like structures observed in cells transfected with pNS2-V5 [23]. …”

Section: Discussionmentioning

confidence: 99%

“…Structural studies of NS2 indicate intermolecular interactions and suggest that it forms helical oligomers that have enhanced stability when interacting with RNA [21-23]. The carboxy terminus of the protein appears to play a role in the assembly of oligomers and therefore the aggregation of NS2 into larger structures [22,23].…”

Section: Introductionmentioning

confidence: 99%

“…The carboxy terminus of the protein appears to play a role in the assembly of oligomers and therefore the aggregation of NS2 into larger structures [22,23]. Phosphorylation reduces the affinity of NS2 for ssRNA and appears to increase the number of VIBs that form near the nucleus, in contrast to the diffuse NS2 staining pattern of the dephosphorylated protein [21].…”

Section: Introductionmentioning

confidence: 99%

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Bluetongue virus infection induces aberrant mitosis in mammalian cells

Shaw

Brüning-Richardson

Morrison

et al. 2013

Virol J

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BackgroundBluetongue virus (BTV) is an arbovirus that is responsible for ‘bluetongue’, an economically important disease of livestock. Although BTV is well characterised at the protein level, less is known regarding its interaction with host cells. During studies of virus inclusion body formation we observed what appeared to be a large proportion of cells in mitosis. Although the modulation of the cell cycle is well established for many viruses, this was a novel observation for BTV. We therefore undertook a study to reveal in more depth the impact of BTV upon cell division.MethodsWe used a confocal microscopy approach to investigate the localisation of BTV proteins in a cellular context with their respective position relative to cellular proteins. In addition, to quantitatively assess the frequency of aberrant mitosis induction by the viral non-structural protein (NS) 2 we utilised live cell imaging to monitor HeLa-mCherry tubulin cells transfected with a plasmid expressing NS2.ResultsOur data showed that these ‘aberrant mitoses’ can be induced in multiple cell types and by different strains of BTV. Further study confirmed multiplication of the centrosomes, each resulting in a separate mitotic spindle during mitosis. Interestingly, the BTV NS1 protein was strongly localised to the centrosomal regions. In a separate, yet related observation, the BTV NS2 protein was co-localised with the condensed chromosomes to a region suggestive of the kinetochore. Live cell imaging revealed that expression of an EGFP-NS2 fusion protein in HeLa-mCherry tubulin cells also results in mitotic defects.ConclusionsWe hypothesise that NS2 is a microtubule cargo protein that may inadvertently disrupt the interaction of microtubule tips with the kinetochores during mitosis. Furthermore, the BTV NS1 protein was distinctly localised to a region encompassing the centrosome and may therefore be, at least in part, responsible for the disruption of the centrosome as observed in BTV infected mammalian cells.

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“…This represents a novel mechanism of modulating RNA chaperone activity, whereby the oligomeric state of the protein defines its unwinding efficiency. Similar to ARV σNS, mammalian reovirus σNS and bluetongue virus NS2 have also been reported to exist in a range of oligomeric states (63)(64)(65)(66). It is possible that different σNS oligomers may play distinct roles during viral replication.…”

Section: Rna-driven Oligomerisationmentioning

confidence: 99%

Stability of local secondary structure determines selectivity of viral RNA chaperones

Bravo

Borodavka

Barth

et al. 2018

Preprint

View full text Add to dashboard Cite

To maintain genome integrity, segmented double-stranded RNA viruses of the Reoviridae family must accurately select and package a complete set of up to a dozen distinct genomic RNAs. It is thought that the high fidelity segmented genome assembly involves multiple sequence-specific RNA-RNA interactions between single-stranded RNA segment precursors. These are mediated by virus-encoded non-structural proteins with RNA chaperone-like activities, such as rotavirus NSP2 and avian reovirus σNS. Here, we compared the abilities of NSP2 and σNS to mediate sequence-specific interactions between rotavirus genomic segment precursors. Despite their similar activities, NSP2 successfully promotes intersegment association, while σNS fails to do so. To understand the mechanisms underlying such selectivity in promoting inter-molecular duplex formation, we compared RNA-binding and helix-unwinding activities of both proteins. We demonstrate that octameric NSP2 binds structured RNAs with high affinity, resulting in efficient intramolecular RNA helix disruption. Hexameric σNS oligomerises into an octamer that binds two RNAs, yet it exhibits only limited RNA-unwinding activity compared to NSP2. Thus, the formation of intersegment RNA-RNA interactions is governed by both helix-unwinding capacity of the chaperones and stability of RNA structure. We propose that this protein-mediated RNA selection mechanism may underpin the high fidelity assembly of multi-segmented RNA genomes in Reoviridae.

show abstract

Structural features of the Bluetongue virus NS2 protein

Cited by 19 publications

References 33 publications

Stability of local secondary structure determines selectivity of viral RNA chaperones

Stability of local secondary structure determines selectivity of viral RNA chaperones

Bluetongue virus infection induces aberrant mitosis in mammalian cells

Stability of local secondary structure determines selectivity of viral RNA chaperones

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