Rewriting nature’s assembly manual for a ssRNA virus

Patel, Nikesh; Wroblewski, Emma; Leonov, German; Phillips, Simon E. V.; Tůma, Roman; Twarock, Reidun; Stockley, Peter G.

doi:10.1073/pnas.1706951114

Cited by 53 publications

(56 citation statements)

References 41 publications

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“…Engineering of AaLS Protein and Its mRNA. Several ssRNA viruses employ genome packaging strategies that rely on direct interactions between arginine-rich motifs (ARMs) on the viral capsid and specific RNA structures in their genome (15)(16)(17). To equip AaLS capsids with ARMs, we relocated the N and C termini of the individual subunits from the exterior to the interior of the cage, between residues 84 and 85, by circular permutation (SI Appendix, Fig.…”

Section: Resultsmentioning

confidence: 99%

Laboratory evolution of virus-like nucleocapsids from nonviral protein cages

Terasaka

Azuma

Hilvert

2018

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

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Viruses are remarkable nanomachines that efficiently hijack cellular functions to replicate and self-assemble their components within a complex biological environment. As all steps of the viral life cycle depend on formation of a protective proteinaceous shell that packages the DNA or RNA genome, bottom-up construction of virus-like nucleocapsids from nonviral materials could provide valuable insights into virion assembly and evolution. Such constructs could also serve as safe alternatives to natural viruses for diverse nano- and biotechnological applications. Here we show that artificial virus-like nucleocapsids can be generated-rapidly and surprisingly easily-by engineering and laboratory evolution of a nonviral protein cage formed by lumazine synthase (AaLS) and its encoding mRNA. Cationic peptides were appended to the engineered capsid proteins to enable specific recognition of packaging signals on cognate mRNAs, and subsequent evolutionary optimization afforded nucleocapsids with expanded spherical structures that encapsulate their own full-length RNA genome in vivo and protect the cargo molecules from nucleases. These findings provide strong experimental support for the hypothesis that subcellular protein-bounded compartments may have facilitated the emergence of ancient viruses.

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Section: Resultsmentioning

confidence: 99%

Laboratory evolution of virus-like nucleocapsids from nonviral protein cages

Terasaka

Azuma

Hilvert

2018

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

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“…This means that 30-segmentation at the second packaging stage is mainly associated with G+C-rich periodic patterns. The significant harmonics in the vicinity of n = 60 in A+T DFT spectrum may be related to AS suggested by Patel et al (2017). The two-stage transition shown in Fig.…”

Section: Discussionmentioning

confidence: 61%

“…13. Patel et al (2017) suggested for the first assembly stage the motifs AN 2 A. One-to-one correspondence between these motifs and coat proteins implies 60segmentation with A+T-rich patterns. The data by Larson et al (2014) unambiguously assign the strikingly pronounced harmonic at n = 30 (G+C, DFT) in Fig.…”

Section: Discussionmentioning

confidence: 94%

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Genome packaging within icosahedral capsids and large-scale segmentation in viral genomic sequences

Chechetkin

Lobzin

2018

Journal of Biomolecular Structure and Dynamics

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The assembly and maturation of viruses with icosahedral capsids must be coordinated with icosahedral symmetry. The icosahedral symmetry imposes also the restrictions on the cooperative specific interactions between genomic RNA/DNA and coat proteins that should be reflected in quasi-regular segmentation of viral genomic sequences. Combining discrete direct and double Fourier transforms, we studied the quasi-regular large-scale segmentation in genomic sequences of different ssRNA, ssDNA, and dsDNA viruses. The particular representatives included satellite tobacco mosaic virus (STMV) and the strains of satellite tobacco necrosis virus (STNV), STNV-C, STNV-1, STNV-2, Escherichia phages MS2, ϕX174, α3, and HK97, and Simian virus 40. In all their genomes, we found the significant quasi-regular segmentation of genomic sequences related to the virion assembly and the genome packaging within icosahedral capsid. We also found good correspondence between our results and available cryo-electron microscopy data on capsid structures and genome packaging in these viruses. Fourier analysis of genomic sequences provides the additional insight into mechanisms of hierarchical genome packaging and may be used for verification of the concepts of 3-fold or 5-fold intermediates in virion assembly. The results of sequence analysis should be taken into account at the choice of models and data interpretation. They also may be helpful for the development of antiviral drugs. Communicated by Ramaswamy H. Sarma.

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“…It has become increasingly appreciated in recent years that for single-stranded RNA viruses, the life cycle is in part regulated by the secondary and tertiary structures of their genomes, hence the current high priority to understand protein-(single-stranded) RNA recognition motifs and RNA sequence-specific folding as it occurs in assembled virions (22,(31)(32)(33)(34)(35). Such information may help with efforts to inhibit viral propagation as well as in nanotechnology applications aimed at harnessing either virus or virus-like systems for efficient gene delivery (36).…”

Section: Discussionmentioning

confidence: 99%

A 2.8-Angstrom-Resolution Cryo-Electron Microscopy Structure of Human Parechovirus 3 in Complex with Fab from a Neutralizing Antibody

Domanska

Flatt

Jukonen

et al. 2019

J Virol

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Human parechovirus 3 (HPeV3) infection is associated with sepsis characterized by significant immune activation and subsequent tissue damage in neonates. Strategies to limit infection have been unsuccessful due to inadequate molecular diagnostic tools for early detection and the lack of a vaccine or specific antiviral therapy. Toward the latter, we present a 2.8-Å-resolution structure of HPeV3 in complex with fragments from a neutralizing human monoclonal antibody, AT12-015, using cryo-electron microscopy (cryo-EM) and image reconstruction. Modeling revealed that the epitope extends across neighboring asymmetric units with contributions from capsid proteins VP0, VP1, and VP3. Antibody decoration was found to block binding of HPeV3 to cultured cells. Additionally, at high resolution, it was possible to model a stretch of RNA inside the virion and, from this, identify the key features that drive and stabilize protein-RNA association during assembly. IMPORTANCE Human parechovirus 3 (HPeV3) is receiving increasing attention as a prevalent cause of sepsis-like symptoms in neonates, for which, despite the severity of disease, there are no effective treatments available. Structural and molecular insights into virus neutralization are urgently needed, especially as clinical cases are on the rise. Toward this goal, we present the first structure of HPeV3 in complex with fragments from a neutralizing monoclonal antibody. At high resolution, it was possible to precisely define the epitope that, when targeted, prevents virions from binding to cells. Such an atomic-level description is useful for understanding host-pathogen interactions and viral pathogenesis mechanisms and for finding potential cures for infection and disease.

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Rewriting nature’s assembly manual for a ssRNA virus

Cited by 53 publications

References 41 publications

Laboratory evolution of virus-like nucleocapsids from nonviral protein cages

Laboratory evolution of virus-like nucleocapsids from nonviral protein cages

Genome packaging within icosahedral capsids and large-scale segmentation in viral genomic sequences

A 2.8-Angstrom-Resolution Cryo-Electron Microscopy Structure of Human Parechovirus 3 in Complex with Fab from a Neutralizing Antibody

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