The fine structure of the DNA-containing core of vaccinia virus

Peters, D.; Müller, G.

doi:10.1016/0042-6822(63)90267-8

Cited by 52 publications

(20 citation statements)

References 8 publications

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“…1). Similar to previous work (Harris and Westwood, 1964; Muller and Peters, 1963; Peters and Muller, 1963), when virions were negative-stained with phospho-tungstic acid (PTA) at pH 7.0 particles could be visualized in two different forms, the mulberry-like form and the capsule (Fig. 1A).…”

Section: Resultssupporting

confidence: 86%

“…Electron microscopy is the preferred method for studying the morphology of the VACV particle and various electron microscopic techniques have been applied in the visualization of the virus structure (Cyrklaff et al, 2005; Dales and Siminovith, 1961; Easterbrook, 1966; Harris and Westwood, 1964; Ichihashi et al, 1984; Muller and Peters, 1963; Naginton and Horne, 1962; Peters and Muller, 1963; Westwood et al, 1964; Wilton et al, 1995). Overall, poxvirus virions have an ellipsoidal, barrel or brick shaped appearance.…”

Section: Introductionmentioning

confidence: 99%

“…Overall, poxvirus virions have an ellipsoidal, barrel or brick shaped appearance. Analysis of VACV on a whole mount preparation using negative staining of the particles revealed the presence of a membrane that enclosed two distinct virus sub-domains: the lateral bodies and the core (Dales, 1962; Harris and Westwood, 1964; Muller and Peters, 1963; Peters and Muller, 1963; Westwood et al, 1964). The lateral bodies, which flank the core, are amorphous structures composed of proteins of unknown function.…”

Section: Introductionmentioning

confidence: 99%

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Fine structure of the vaccinia virion determined by controlled degradation and immunolocalization

Moussatché

Condit

2015

Virology

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The vaccinia virion is a membraned, slightly flattened, barrel-shaped particle, with a complex internal structure featuring a biconcave core flanked by lateral bodies. Although the architecture of the purified mature virion has been intensely characterized by electron microscopy, the distribution of the proteins within the virion has been examined primarily using biochemical procedures. Thus, it has been shown that non-ionic and ionic detergents combined or not with a sulfhydryl reagent can be used to disrupt virions and, to a limited degree, separate the constituent proteins in different fractions. Applying a controlled degradation technique to virions adsorbed on EM grids, we were able to immuno-localize viral proteins within the virion particle. Our results show after NP40 and DTT treatment, membrane proteins are removed from the virion surface revealing proteins that are associated with the lateral bodies and the outer layer of the core wall. Combined treatment using high salt and high DTT removed lateral body proteins and exposed proteins of the internal core wall. Cores treated with proteases could be disrupted and the internal components were exposed. Cts8, a mutant in the A3 protein, produces aberrant virus that, when treated with NP-40 and DTT, release to the exterior the virus DNA associated with other internal core proteins. With these results, we are able to propose a model for the structure the vaccinia virion.

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Section: Resultssupporting

confidence: 86%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Fine structure of the vaccinia virion determined by controlled degradation and immunolocalization

Moussatché

Condit

2015

Virology

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“…However, the complex features on the bottom and inside of the particle were not revealed, and perhaps such a clear, and for its time excellent, model helped to reinforce the idea that vaccinia virus has a rather symmetrical, brick-shaped structure. In later papers (39,40) this author revealed unexpected complexity in the structure of a triad of bulbous elements that seemed to be made up of highly coiled tubules. This was also the interpretation of Hohenberg, who made a detailed artistic drawing of the virus (see reference 34).…”

Section: Discussionmentioning

confidence: 99%

Structure and Assembly of Intracellular Mature Vaccinia Virus: Isolated-Particle Analysis

Griffiths

Wepf

Wendt

et al. 2001

J Virol

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In a series of papers, we have provided evidence that during its assembly vaccinia virus is enveloped by a membrane cisterna that originates from a specialized, virally modified, smooth-membraned domain of the endoplasmic reticulum ( In this first study, we analyzed the IMV by on-grid staining, cryoscanning EM (SEM), and cryo-transmission EM. We focused on the structure of the IMV particle, both after isolation and in the context of viral entry. For the latter, we used high-resolution cryo-SEM combined with cryofixation, as well as a novel approach we developed for investigating vaccinia IMV bound to plasma membrane fragments adsorbed onto EM grids. Our analysis revealed that the IMV is made up of interconnected cisternal and tubular domains that fold upon themselves via a complex topology that includes an S-shaped fold. The viral tubules appear to be eviscerated from the particle during viral infection. Since the structure of the IMV is the result of a complex assembly process, we also provide a working model to explain how a specialized smooth-ER domain can be modulated to form the IMV. We also present theoretical arguments for why it is highly unlikely that the IMV is surrounded by only a single membrane.

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“…Experiments from the early 1960s revealed a tubular structure inside the core (6,7). This tubular structure has been largely overlooked for the last 50 years because of the difficulty in preserving it using conventional techniques for electron microscopy (EM).…”

mentioning

confidence: 99%

Vaccinia Virus Mutations in the L4R Gene Encoding a Virion Structural Protein Produce Abnormal Mature Particles Lacking a Nucleocapsid

Jesus

Moussatché

Condit

2014

J Virol

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Electron micrographs from the 1960s revealed the presence of an S-shaped tubular structure in the center of the vaccinia virion core. Recently, we showed that packaging of virus transcription enzymes is necessary for the formation of the tubular structure, suggesting that the structure is equivalent to a nucleocapsid. Based on this study and on what is known about nucleocapsids of other viruses, we hypothesized that in addition to transcription enzymes, the tubular structure also contains the viral DNA and a structural protein as a scaffold. The vaccinia virion structural protein L4 stands out as the best candidate for the role of a nucleocapsid structural protein because it is abundant, it is localized in the center of the virion core, and it binds DNA. In order to gain more insight into the structure and relevance of the nucleocapsid, we analyzed thermosensitive and inducible mutants in the L4R gene. Using a cryo-fixation method for electron microscopy (high-pressure freezing followed by freeze-substitution) to preserve labile structures like the nucleocapsid, we were able to demonstrate that in the absence of functional L4, mature particles with defective internal structures are produced under nonpermissive conditions. These particles do not contain a nucleocapsid. In addition, the core wall of these virions is abnormal. This suggests that the nucleocapsid interacts with the core wall and that the nucleocapsid structure might be more complex than originally assumed. IMPORTANCEThe vaccinia virus nucleocapsid has been neglected since the 1960s due to a lack of electron microscopy techniques to preserve this labile structure. With the advent of cryo-fixation techniques, like high-pressure freezing/freeze-substitution, we are now able to consistently preserve and visualize the nucleocapsid. Because vaccinia virus early transcription is coupled to the viral core structure, detailing the structure of the nucleocapsid is indispensable for determining the mechanisms of vaccinia virus core-directed transcription. The present study represents our second attempt to understand the structure and biological significance of the nucleocapsid. We demonstrate the importance of the protein L4 for the formation of the nucleocapsid and reveal in addition that the nucleocapsid and the core wall may be associated, suggesting a higher level of complexity of the nucleocapsid than predicted. In addition, we prove the utility of high-pressure freezing in preserving the vaccinia virus nucleocapsid.

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The fine structure of the DNA-containing core of vaccinia virus

Cited by 52 publications

References 8 publications

Fine structure of the vaccinia virion determined by controlled degradation and immunolocalization

Fine structure of the vaccinia virion determined by controlled degradation and immunolocalization

Structure and Assembly of Intracellular Mature Vaccinia Virus: Isolated-Particle Analysis

Vaccinia Virus Mutations in the L4R Gene Encoding a Virion Structural Protein Produce Abnormal Mature Particles Lacking a Nucleocapsid

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