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Bacteriophage MS2 genomic RNA encodes an assembly instruction manual for its capsid.

Stockley PG, White SJ, Dykeman E, Manfield I, Rolfsson O, Patel N, Bingham R, Barker A, Wroblewski E, Chandler-Bostock R, Weiß EU, Ranson NA, Tuma R, Twarock R - Bacteriophage (2016)

Bottom Line: They act co-operatively to regulate the dominant assembly pathway and ensure cognate RNA encapsidation.In MS2, they also trigger conformational change in the dimeric capsomere creating the A/B quasi-conformer, 60 of which are needed to complete the T=3 capsid.This is the most compelling demonstration to date that this ssRNA virus, and by implications potentially very many of them, assemble via a PS-mediated assembly mechanism.

View Article: PubMed Central - PubMed

Affiliation: Astbury Center for Structural Molecular Biology, University of Leeds , Leeds, UK.

ABSTRACT

Using RNA-coat protein crosslinking we have shown that the principal RNA recognition surface on the interior of infectious MS2 virions overlaps with the known peptides that bind the high affinity translational operator, TR, within the phage genome. The data also reveal the sequences of genomic fragments in contact with the coat protein shell. These show remarkable overlap with previous predictions based on the hypothesis that virion assembly is mediated by multiple sequences-specific contacts at RNA sites termed Packaging Signals (PSs). These PSs are variations on the TR stem-loop sequence and secondary structure. They act co-operatively to regulate the dominant assembly pathway and ensure cognate RNA encapsidation. In MS2, they also trigger conformational change in the dimeric capsomere creating the A/B quasi-conformer, 60 of which are needed to complete the T=3 capsid. This is the most compelling demonstration to date that this ssRNA virus, and by implications potentially very many of them, assemble via a PS-mediated assembly mechanism.

No MeSH data available.


Related in: MedlinePlus

The implications of PS-mediated assembly for the earliest stages of infection. Shown is the asymmetric tomographic structure of MS2, at low resolution, bound to its initial cellular receptor, the E.coli F-pilus. This structure shows extensive density for the viral RNA, confirming the idea that PS-mediated assembly leads to almost identical RNA conformations within each viral particle. The contact to the pilus is mediated by the maturation protein, MP, which also binds to specific sites close to either end of the viral genome. Only the MP-RNA complex enters the bacterial cell, leaving the CP shell largely intact. MP replaces a C/C capsomere in an otherwise icosahedral protein shell.
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f0003: The implications of PS-mediated assembly for the earliest stages of infection. Shown is the asymmetric tomographic structure of MS2, at low resolution, bound to its initial cellular receptor, the E.coli F-pilus. This structure shows extensive density for the viral RNA, confirming the idea that PS-mediated assembly leads to almost identical RNA conformations within each viral particle. The contact to the pilus is mediated by the maturation protein, MP, which also binds to specific sites close to either end of the viral genome. Only the MP-RNA complex enters the bacterial cell, leaving the CP shell largely intact. MP replaces a C/C capsomere in an otherwise icosahedral protein shell.

Mentions: Virions are necessarily transient structures, acting as transport vehicles for the genome between hosts. In a large number of cases the initial steps in infection for positive-sense, ssRNA viruses involve contact with a cellular receptor leading to conformational changes within the viral shell ultimately enabling the extrusion of the genome through a unique capsomer or vertex. For such an infection mechanism to occur the virus must be able to position one end of its RNA close to or beneath that special vertex. PS-mediated assembly can provide that control. In the case of MS2 there is an additional factor, namely a single copy maturation protein (MP) that both serves as the attachment point to the initial bacterial receptor, the pilus (Fig. 3) and guides the RNA-MP complex, formed via distinct MP PSs, into the target cell. Mathematical modeling of such an assembly reaction suggests that it ensures cognate and complete capsid assembly via the most stable assembly pathways,11 and that this can only be observed if the CP concentration is slowly ramped up as in a real infection. Perhaps this is one reason why PS-mediated assembly has been overlooked in previous in vitro reassembly reactions, while it was unambiguously demonstrated by us in the presence of such a ramp.12 In effect, the RNA forms a unique sequence of RNA-CP contacts with the overlying CP shell, a problem akin to the dilemma faced by a traveling salesman, i.e. identification of the best route to visit a number of defined sites once and only once while avoiding doubling back. These results imply an astonishing prediction, namely that there are only a very limited number of conformations of the RNA in proximity to the CP layer within every viral particle. Indeed, the interpretation of the recent cross-linking data assumes this to be the case.Figure 3.


Bacteriophage MS2 genomic RNA encodes an assembly instruction manual for its capsid.

Stockley PG, White SJ, Dykeman E, Manfield I, Rolfsson O, Patel N, Bingham R, Barker A, Wroblewski E, Chandler-Bostock R, Weiß EU, Ranson NA, Tuma R, Twarock R - Bacteriophage (2016)

The implications of PS-mediated assembly for the earliest stages of infection. Shown is the asymmetric tomographic structure of MS2, at low resolution, bound to its initial cellular receptor, the E.coli F-pilus. This structure shows extensive density for the viral RNA, confirming the idea that PS-mediated assembly leads to almost identical RNA conformations within each viral particle. The contact to the pilus is mediated by the maturation protein, MP, which also binds to specific sites close to either end of the viral genome. Only the MP-RNA complex enters the bacterial cell, leaving the CP shell largely intact. MP replaces a C/C capsomere in an otherwise icosahedral protein shell.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4836477&req=5

f0003: The implications of PS-mediated assembly for the earliest stages of infection. Shown is the asymmetric tomographic structure of MS2, at low resolution, bound to its initial cellular receptor, the E.coli F-pilus. This structure shows extensive density for the viral RNA, confirming the idea that PS-mediated assembly leads to almost identical RNA conformations within each viral particle. The contact to the pilus is mediated by the maturation protein, MP, which also binds to specific sites close to either end of the viral genome. Only the MP-RNA complex enters the bacterial cell, leaving the CP shell largely intact. MP replaces a C/C capsomere in an otherwise icosahedral protein shell.
Mentions: Virions are necessarily transient structures, acting as transport vehicles for the genome between hosts. In a large number of cases the initial steps in infection for positive-sense, ssRNA viruses involve contact with a cellular receptor leading to conformational changes within the viral shell ultimately enabling the extrusion of the genome through a unique capsomer or vertex. For such an infection mechanism to occur the virus must be able to position one end of its RNA close to or beneath that special vertex. PS-mediated assembly can provide that control. In the case of MS2 there is an additional factor, namely a single copy maturation protein (MP) that both serves as the attachment point to the initial bacterial receptor, the pilus (Fig. 3) and guides the RNA-MP complex, formed via distinct MP PSs, into the target cell. Mathematical modeling of such an assembly reaction suggests that it ensures cognate and complete capsid assembly via the most stable assembly pathways,11 and that this can only be observed if the CP concentration is slowly ramped up as in a real infection. Perhaps this is one reason why PS-mediated assembly has been overlooked in previous in vitro reassembly reactions, while it was unambiguously demonstrated by us in the presence of such a ramp.12 In effect, the RNA forms a unique sequence of RNA-CP contacts with the overlying CP shell, a problem akin to the dilemma faced by a traveling salesman, i.e. identification of the best route to visit a number of defined sites once and only once while avoiding doubling back. These results imply an astonishing prediction, namely that there are only a very limited number of conformations of the RNA in proximity to the CP layer within every viral particle. Indeed, the interpretation of the recent cross-linking data assumes this to be the case.Figure 3.

Bottom Line: They act co-operatively to regulate the dominant assembly pathway and ensure cognate RNA encapsidation.In MS2, they also trigger conformational change in the dimeric capsomere creating the A/B quasi-conformer, 60 of which are needed to complete the T=3 capsid.This is the most compelling demonstration to date that this ssRNA virus, and by implications potentially very many of them, assemble via a PS-mediated assembly mechanism.

View Article: PubMed Central - PubMed

Affiliation: Astbury Center for Structural Molecular Biology, University of Leeds , Leeds, UK.

ABSTRACT

Using RNA-coat protein crosslinking we have shown that the principal RNA recognition surface on the interior of infectious MS2 virions overlaps with the known peptides that bind the high affinity translational operator, TR, within the phage genome. The data also reveal the sequences of genomic fragments in contact with the coat protein shell. These show remarkable overlap with previous predictions based on the hypothesis that virion assembly is mediated by multiple sequences-specific contacts at RNA sites termed Packaging Signals (PSs). These PSs are variations on the TR stem-loop sequence and secondary structure. They act co-operatively to regulate the dominant assembly pathway and ensure cognate RNA encapsidation. In MS2, they also trigger conformational change in the dimeric capsomere creating the A/B quasi-conformer, 60 of which are needed to complete the T=3 capsid. This is the most compelling demonstration to date that this ssRNA virus, and by implications potentially very many of them, assemble via a PS-mediated assembly mechanism.

No MeSH data available.


Related in: MedlinePlus