Cauliflower mosaic virus: A 420 subunit (T = 7), multilayer structure

Abstract

The structures of the Cabb-B and CM 1841 strains of cauliflower mosaic virus (CaMV) have been solved to about 3 nm resolution from unstained, frozen-hydrated samples that were examined with low-irradiation cryo-electron microscopy and three-dimensional image reconstruction procedures. CaMV is highly susceptible to distortions. Spherical particles, with a maximum diameter of 53.8 nm, are composed of three concentric layers (I-III) of solvent-excluded density that surround a large, solvent-filled cavity (∼27 nm dia.). The outermost layer (I) contains 72 capsomeric morphological units, with 12 pentavalent pentamers and 60 hexavalent hexamers for a total of 420 subunits (37-42 kDa each) arranged with T = 7 icosahedral symmetry. CaMV is the first example of a T = 7 virus that obeys the rules of stoichiometry proposed for isometric viruses by Caspar and Klug (1962, Cold Spring Harb. Symp. Quant. Biol. 27, 1-24), although the hexameric capsomers exhibit marked departure from the regular sixfold symmetry expected for a structure in which the capsid protein subunits are quasi-equivalently related. The double-stranded DNA genome is distributed in layers II and III along with a portion of the viral protein. The CaMV reconstructions are consistent with the model based on neutron diffraction studies (Kruse et al., 1987, Virology 159, 166-168) and, together, these structural models are discussed in relation to a replicationassembly model (Hull et al., 1987, J. Cell Sci. (Suppl.) 7, 213-229). Remarkable agreement between the reconstructions of CaMV Cabb-B and CM1841 suggests that other strains of CaMV adopt the Same basic Structure.

A Model-Based Approach for Determining Orientations of Biological Macromolecules Imaged by Cryoelectron Microscopy

“…Common Lines often fails or leads to inconsistent results with cryoEM images recorded close to focus (<1.5 m) or with viruses that do not have coarse morphological features (see Example 4, below). Our Common Lines analysis of CaMV only succeeded after we recorded focal pairs of vitrified specimens and used a 2.4-m underfocus micrograph to determine initial orientations of the particles and applied these values to the same particles in the closer-to-focus (1.2-m), lower-dose image (Cheng et al, 1992). As a test, we reexamined the same 1.2-m image with Common Lines and were only able to identify the correct orientation view for 1 of the 21 CaMV particle images that went into the original 3D reconstruction.…”

“…At first glance these two viruses appear quite similar with prominent ''caps'' of density at the 12 icosahedral vertices. Closer inspection reveals many striking differences in the capsid sub- , and 7 of the refinement: the refined models and the original, 21-particle reconstruction of CaMV obtained by Common Lines (Cheng et al, 1992) (Griffith et al, 1992). structure.…”

“…4a2-4a8) show how they converge to a final and correct solution. The starting model contains only very-low-resolution information (spheres) yet the final reconstruction shows 12 capsomers with pentameric substructure and 60 capsomers with hexameric substructure as was discovered in the original analysis of this structure (Cheng et al, 1992). Correlation coefficients computed in reciprocal ( Fig.…”

Ab initio random model method facilitates 3D reconstruction of icosahedral particles

“…Wikoff, et al, 1994), or a simple, geometric structure constructed in silico (e.g. Cheng, et al, 1992). The potential for model-induced bias in the reconstruction process always exists with indirect models since they are not derived from the images being analyzed.…”

Structural Insights into the Molecular Mechanisms of Cauliflower Mosaic Virus Transmission by Its Insect Vector

“…It is constituted of three concentric shells, built from 420 capsid protein subunits, surrounding a large inner cavity with a diameter of approximately 250 Å (4,37). The physical association between P3 and viral particles has been consistently demonstrated in mature CaMV virions by copurification, immunolabeling experiments, and in vitro interactions (7,(23)(24)(25).…”