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

Stockley, Peter G.; White, Simon; Dykeman, Eric C.; Manfield, Iain; Rolfsson, Óttar; Patel, Nikesh; Bingham, Richard J.; Barker, Amy; Wroblewski, Emma; Chandler-Bostock, Rebecca; Weiß, Eva U.; Ranson, Neil A.; Tůma, Roman; Twarock, Reidun

doi:10.1080/21597081.2016.1157666

Cited by 40 publications

(44 citation statements)

References 38 publications

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“…MS2 infects E. coli by interacting with its F‐pilus through the maturation protein (Valentine and Strand, ; Brinton, Gemski, and Carnahan, ). This interaction allows the phage to deliver its genome and the MP inside the host cell, while the capsid shell is emptied and left outside the bacterium (Stockley et al , ). Therefore, viral particles lacking MP lose infectivity and do not bind to E. coli F‐pili (Krahn et al , ; Roberts and Steitz, ).…”

Section: Introductionmentioning

confidence: 99%

“…MS2 particles in vivo have capsids with T = 3 pseudo‐icosahedral symmetry (Valegard et al , ; Golmohammadi et al , ) that are co‐assembled around the genome. The ssRNA forms secondary structural elements that directly interact with the coat proteins acting as packaging signals, with the 19‐nt long stem loop being the best characterised (Stockley et al , ). During the assembly process, the RNA specifies the three quasi‐equivalent conformations of the coat protein that form the IAU, named A, B and C (Stockley et al , ).…”

Section: Introductionmentioning

confidence: 99%

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Bacteriophage MS2 displays unreported capsid variability assembling T = 4 and mixed capsids

Garrido

Crone

Ramlaul

et al. 2019

Molecular Microbiology

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Bacteriophage MS2 is a positive-sense, singlestranded RNA virus encapsulated in an asymmetric T = 3 pseudo-icosahedral capsid. It infects Escherichia coli through the F-pilus, in which it binds through a maturation protein incorporated into its capsid. Cryogenic electron microscopy has previously shown that its genome is highly ordered within virions, and that it regulates the assembly process of the capsid. In this study, we have assembled recombinant MS2 capsids with non-genomic RNA containing the capsid incorporation sequence, and investigated the structures formed, revealing that T = 3, T = 4 and mixed capsids between these two triangulation numbers are generated, and resolving structures of T = 3 and T = 4 capsids to 4 Å and 6 Å respectively. We conclude that the basic MS2 capsid can form a mix of T = 3 and T = 4 structures, supporting a role for the ordered genome in favouring the formation of functional T = 3 virions.Abbreviations: CP, coat protein; IAU, icosahedral asymmetric unit; MP, maturation protein; VLP, virus-like particle.

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Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Bacteriophage MS2 displays unreported capsid variability assembling T = 4 and mixed capsids

Garrido

Crone

Ramlaul

et al. 2019

Molecular Microbiology

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“…Once thought to be noncatalytic messengers and scaffolds for use in translation, RNA has been shown to play active roles in such diverse processes as mRNA splicing (Kruger et al 1982;McNeil et al 2016), regulation of transcription (Grundy et al 1994;Henkin 1994;Serganov and Nudler 2013;Furtig et al 2015) and translation (Lee et al 1993;Wightman et al 1993;Filipowicz et al 2008), viral assembly (Zeffman et al 2000;Cantara et al 2014;Sardo et al 2015;Stockley et al 2016), and immunity (Brouns et al 2008;Zhang et al 2010;Dhahbi 2015;Cavalieri et al 2016). Many of these functions result from the ability of RNA to fold into complex secondary and tertiary structures.…”

Section: Introductionmentioning

confidence: 99%

RiboCAT: a new capillary electrophoresis data analysis tool for nucleic acid probing

Cantara

Hatterschide

et al. 2016

RNA

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Chemical and enzymatic probing of RNA secondary structure and RNA/protein interactions provides the basis for understanding the functions of structured RNAs. However, the ability to rapidly perform such experiments using capillary electrophoresis has been hampered by relatively labor-intensive data analysis software. While these computationally robust programs have been shown to calculate residue-specific reactivities to a high degree of accuracy, they often require time-consuming manual intervention and lack the ability to be easily modified by users. To alleviate these issues, RiboCAT (Ribonucleic acid capillaryelectrophoresis analysis tool) was developed as a user-friendly, Microsoft Excel-based tool that reduces the need for manual intervention, thereby significantly shortening the time required for data analysis. Features of this tool include (i) the use of an Excel platform, (ii) a method of intercapillary signal alignment using internal size standards, (iii) a peak-sharpening algorithm to more accurately identify peaks, and (iv) an open architecture allowing for simple user intervention. Furthermore, a complementary tool, RiboDOG (RiboCAT data output generator) was designed to facilitate the comparison of multiple data sets, highlighting potential inconsistencies and inaccuracies that may have occurred during analysis. Using these new tools, the secondary structure of the HIV-1 5 ′ untranslated region (5 ′ UTR) was determined using selective 2 ′ -hydroxyl acylation analyzed by primer extension (SHAPE), matching the results of previous work.

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“…Unlike most dsDNA phages, which use specialized protein machinery to pump their genomic DNA into a capsid preassembled around a protein scaffold (14)(15)(16)(17), ssRNA viruses, including the Leviviridae, assemble their coat proteins around the genomic RNA (gRNA), presumably because the extremely small genome does not allow for genes to encode proteins that help package the genetic material. Therefore, ssRNA viruses require direct gRNA-coat protein interactions, some of which are specific, to self-assemble the virion (18). This raises interesting questions, including how the viral RNA is selectively encapsidated over host RNAs.…”

mentioning

confidence: 99%

Asymmetric cryo-EM structure of the canonical Allolevivirus Qβ reveals a single maturation protein and the genomic ssRNA in situ

Gorzelnik

Zhao

Reed

et al. 2016

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

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Single-stranded (ss) RNA viruses infect all domains of life. To date, for most ssRNA virions, only the structures of the capsids and their associated protein components have been resolved to high resolution. Qβ, an ssRNA phage specific for the conjugative F-pilus, has a T = 3 icosahedral lattice of coat proteins assembled around its 4,217 nucleotides of genomic RNA (gRNA). In the mature virion, the maturation protein, A 2 , binds to the gRNA and is required for adsorption to the F-pilus. Here, we report the cryo-electron microscopy (cryo-EM) structures of Qβ with and without symmetry applied. The icosahedral structure, at 3.7-Å resolution, resolves loops not previously seen in the published X-ray structure, whereas the asymmetric structure, at 7-Å resolution, reveals A 2 and the gRNA. A 2 contains a bundle of α-helices and replaces one dimer of coat proteins at a twofold axis. The helix bundle binds gRNA, causing denser packing of RNA in its proximity, which asymmetrically expands the surrounding coat protein shell to potentially facilitate RNA release during infection. We observe a fixed pattern of gRNA organization among all viral particles, with the major and minor grooves of RNA helices clearly visible. A single layer of RNA directly contacts every copy of the coat protein, with one-third of the interactions occurring at operator-like RNA hairpins. These RNA-coat interactions stabilize the tertiary structure of gRNA within the virion, which could further provide a roadmap for capsid assembly.single-particle cryo-EM | Allolevivirus | ssRNA virus | genomic RNA | maturation protein S ingle-stranded (ss) RNA viruses are an abundant type of virus and infect all domains of life (1-4). One of the beststudied ssRNA virus systems is the Leviviridae, which infects Gram-negative bacteria via a variety of retractile pili (5); extensive genetic and biochemical studies have been performed on two of these phages: MS2 and Qβ (5-11). All of the Leviviridae have the same core genome, spanning 3.4-4.3 kb, encoding the maturation protein, the coat protein, and a subunit of RNAdependent RNA replicase (SI Appendix, Fig. S1) (10). The MS2-like phages, designated true leviviruses, have a fourth gene that encodes the lysis protein, whereas the Qβ-like phages, designated alloleviviruses, have the lysis function as an additional feature of the maturation protein (called A 2 in Qβ). Qβ also encodes a minor coat protein, called A 1 , arising from occasional readthrough of the stop codon of the major coat protein; it has been estimated that the A 1 protein replaces 3-10 copies of the major coat protein in the virion (11) and is required for infection (12). A 1 consists of a coat domain and a read-through domain separated by a flexible linker (13). Unlike most dsDNA phages, which use specialized protein machinery to pump their genomic DNA into a capsid preassembled around a protein scaffold (14-17), ssRNA viruses, including the Leviviridae, assemble their coat proteins around the genomic RNA (gRNA), presumably because the extremely small ...

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

Cited by 40 publications

References 38 publications

Bacteriophage MS2 displays unreported capsid variability assembling T = 4 and mixed capsids

Bacteriophage MS2 displays unreported capsid variability assembling T = 4 and mixed capsids

RiboCAT: a new capillary electrophoresis data analysis tool for nucleic acid probing

Asymmetric cryo-EM structure of the canonical Allolevivirus Qβ reveals a single maturation protein and the genomic ssRNA in situ

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