Species: Turnip yellow mosaic virus
|This is a revised version of DPV 2|
R. E. F. Matthews
Department of Cell Biology, University of Auckland, New Zealand
Host Range and Symptomatology
Transmission by Vectors
Transmission through Seed
Transmission by Grafting
Transmission by Dodder
Nucleic Acid Hybridization
Stability in Sap
Properties of Particles
Properties of Infective Nucleic Acid
Relations with Cells and Tissues
Ecology and Control
A virus with RNA-containing icosahedral particles 28 nm in diameter. Host range is confined almost entirely to the Cruciferae. Infected cells show a characteristic rounding and clumping of the chloroplasts, readily observed by light microscopy. The virus may reach high concentrations in infected leaves. It is readily transmissible by sap inoculation, and is transmitted in the field by flea-beetles. It has been reported from several European countries.
The Northumberland isolate (Broadbent & Heathcote, 1958). This isolate is serologically distinguishable from the Cambridge culture, and causes less severe symptoms in most hosts, but much more severe ones in cabbage, cauliflower and Brussels sprout.
Other strains collected in the field. Symons et al. (1963) studied the RNA and protein composition of six isolates obtained from various sources in England, Germany and Denmark. They fell clearly into two groups based on their cytosine contents (approximately 38%, or 42%). Analyses of the coat protein indicated a similar grouping.
Four strains were recognized among isolates collected in the German Democratic Republic (Shukla & Schmelzer, 1974). These could be distinguished both serologically and by symptoms on test plants.
(i) Empty protein shells, called T (for top component).
(ii) Infective nucleoproteins (B or bottom components) and particles derived from them. B component separates in CsCl gradients into components B1a and B1b, which are equally infective (Matthews, 1974), and a third component B1c, only recently characterized, which is more dense than B1b. B1b and B1c both contain copies of the coat protein mRNA as well as a molecule of genome RNA (C. W. A. Pleij, personal communication). B1 components can be converted in strong solutions of CsCl, especially at pH > 6.5, to a B2 series with higher densities, designated B2a, B2b and B2c. Their formation is prevented by the presence of 0.1 M MgCl2 in the CsCl (C. W. A. Pleij, personal communication).
(iii) Non-infective nucleoprotein particles containing subgenomic RNA species and having densities in CsCl intermediate between those of the T and B components. Matthews (1960) described three such components. Keeling et al. (1979), using an improved density gradient, resolved eight as illustrated in Fig.7. These are numbered 1-8 in order of increasing RNA content. C. W. A. Pleij (personal communication) has now isolated two further components; the numbering of the components must therefore remain flexible.
The properties listed below for B1 were determined on a mixture of B1a, B1b and (presumably) B1c.
Sedimentation coefficients (s20,w) at infinite dilution: 53-54 (T); 116-117 (B1).
Molecular weights: 3.6 x 106 (T); 5.4 x 106 (B1).
Diffusion coefficients (D20 x 10-7 cm2 sec-1); 1.51 (T), 1.55 (B1).
Isoelectric points: pH 3 .75 (T and B1).
Partial specific volumes: 0.733 (T); 0.661 (B1).
Absorbances at 260 nm (1 mg/ml, 1 cm light path): 0.96 (T); 9.6 (B1).
A260/A280: 0.81 (T); 1.51 (B1).
Buoyant densities in CsCl (g/cm3): 1.28 (T); 1.395 (B1a) and 1.402 (B1b).
Some properties of the minor non-infective nucleoprotein fractions have been determined by
Mellema et al. (1979)
Keeling et al. (1979).
They contain the coat protein
mRNA and one or more of eight other subgenomic RNA species of discrete size (see Particle
Composition) which have not been firmly allocated to particular nucleoprotein species. If all
possible packaging combinations exist there may in fact be a very large number of particles of
slightly differing density. Thus the following data may be subject to revision; the first
figure is the effective buoyant density in CsCl (g/cm3), the second is the ratio
the third and fourth (where available) are the calculated %
RNA and the approximate total weight of RNA in the particle (x 10-6).
Component 1:- 1.294; 1.135; 5%; (mainly coat protein gene).
Component 2:- 1.312; 1.310; 9%; 0.5.
Component 3:- 1.315; 1.286; 11%; 0.6.
Component 4:- 1.330; 1.325.
Component 5:- 1.340; 1.303; 17%; 0.9.
Component 6:- 1.345; 1.452.
Component 7:- 1.357; 1.504; 25%; 1.4.
Component 8:- 1.363; 1.507; 28%; 16.
The proportion of total minor nucleoproteins relative to the infective nucleoproteins is about 5% on a particle number basis.
Among the icosahedral viruses, turnip yellow mosaic virus has a relatively stable structure. Protein-protein interactions are strong, as evidenced by the existence of the stable empty protein shells. However when the virus is taken to pH 11.6 in 1 M KCl the particle swells rapidly (radius increases by about 4%) and the RNA escapes, leaving an empty shell (Kaper, 1975; Keeling et al., 1979). Escape of the RNA is accompanied by a loss from the shell of an amount of protein equivalent to five protein subunits. The minor nucleoproteins do not swell or lose RNA or protein under the same conditions (J. Keeling & R. E. F. Matthews, unpublished data).
Protein: About 65% of the virus by weight. There is probably only one kind of protein in the icosahedral shell. This has a M. Wt of 20,133, and contains 189 amino acid residues of known sequence (Peter et al., 1972).
Other components: Polyamines, mainly spermidine, make up about 1% by weight of the virus (Beer & Kosuge, 1970). No lipid, carbohydrate or enzyme activities are found in the virus.
In the wheat germ system for in vitro protein synthesis the genome RNA is translated to give two large polypeptides with M. Wt c. 180,000 and 150,000 but there is no product corresponding to coat protein. Such a product is, however, translated from the 0.25 x 106 M. Wt RNA; this therefore probably acts in vivo as a messenger for coat protein (Ricard et al., 1977; Pleij et al., 1977; Higgins et al., 1978). The other sub-genomic RNA species each give a polypeptide product in the wheat germ system that represents approximately their full coding capacity; they share a common 5' terminus which may be the same as that of the genome RNA (Mellema et al., 1979) and they probably form a family of readthrough messenger RNAs for polypeptides required for virus synthesis in vivo. This idea is supported by the fact that infection of u.v.-irradiated protoplasts induces the synthesis of a series of polypeptides with sizes that correspond approximately to those expected from the in vitro studies (Y. Sugimura & R. E. F. Matthews, unpublished data).
Electron microscopy reveals a variety of other tissue abnormalities which depend on strain of virus and on the time after infection (Ushiyama & Matthews, 1970). Many of these changes are secondary effects occurring after virus replication has begun. The most constant and characteristic cytological effect of infection is the development of small vesicles near the periphery of the chloroplasts. These vesicles are formed by an invagination of both of the chloroplast bilayer membranes. The vesicle necks are open to the cytoplasm (Fig.5, Fig.6). The double-stranded RNA found in infected tissue, and the virus-induced RNA-dependent RNA polymerase are associated with these vesicles, which are therefore a major site of virus RNA synthesis (Laflèche et al., 1972). They are the earliest consequence of infection detectable by electron microscopy (Hatta & Matthews, 1974) and their presence has been used as a marker of infection to establish a sequence of cytological changes.
Following the appearance of scattered peripheral vesicles, the chloroplasts become rounded. Clusters of closely spaced vesicles can be observed with endoplasmic reticulum lying over them in the cytoplasm (Fig.8). The endoplasmic reticulum is then replaced by electron lucent material which is virus coat protein. Virus particles can be detected first in the cytoplasm overlying the electron lucent material (Fig.9). Virus protein, probably in the form of empty protein shells, accumulates in the nuclei from an early stage after infection (Hatta & Matthews, 1976).
Brassica protoplasts can be infected in vitro. Virus accumulates between about 12 and 36 h after inoculation to give about 106 particles/protoplast (Renaudin et al., 1975). This is only about one tenth of the yield from palisade mesophyll cells in infected leaves. In infected leaves the ratio of empty protein shells /full nucleoprotein particles is usually in the range 0.2 to 0.3. In protoplasts infected in vitro it is close to 1.0 throughout the virus growth cycle.
Mosaic symptoms in Chinese cabbage. Top left is a healthy leaf. The other leaves are from plants inoculated with strains isolated from different islands of tissue in leaves containing the Cambridge stock culture. Top right, pale green; bottom left, yellow green; bottom right, white. (Photo. J. Endt.)
An octahedral crystal of virus particles grown in 0.75 M ammonium sulphate. Bar represents 25 µm. (Photo. T. Hatta.)
Morphological subunits seen in virus particles negatively stained with uranyl acetate. Bar represents 30 nm. (Photo. S. Bullivant.)
Development of rounding and clumping of chloroplasts in a Chinese cabbage mesophyll protoplast inoculated in vitro: (left) 12 h after inoculation, chloroplasts essentially normal; (right) the same protoplast 17 h after inoculation, rounding and clumping of chloroplasts complete. Bar represents 15 µm. (Photo. Y. Sugimura.)
Small peripheral vesicles with necks, induced in Chinese cabbage chloroplasts. Thin section showing continuity of the vesicle membranes with the chloroplast membranes, and stranded material with the staining properties of double-stranded nucleic acid. Bar represents 50 nm. (Photo. S. Bullivant.)
Small peripheral vesicles with necks, induced in Chinese cabbage chloroplasts. Fine structure of vesicle membranes and necks revealed by freeze-fracturing. Bar represents 100 nm. (From Hatta, Bullivant & Matthews, 1973.)
Fractionation of a purified virus preparation into 12 nucleoprotein bands and a band of empty protein shells (T) in a CsCl gradient. Separation of the two infective nucleoprotein bands (B1a and B1b) is obscured by the overloading necessary to reveal some of the minor components (From Keeling et al., 1979.)
Chloroplast in an infected Chinese cabbage cell showing the clustered vesicles at the time they are overlaid by endoplasmic reticulum (ER) in the cytoplasm. Arrow shows a connection between ER and chloroplast membrane. Bar represents 250 nm. (From Hatta & Matthews, 1974.)
Chloroplast in an infected Chinese cabbage cell at a slightly later stage than that in Fig.8. Overlying the electron-lucent material is a small array of virus particles. The tissue was plasmolysed with sucrose before fixation to induce crystallisation of any virus present. Bar represents 250 nm. (From Hatta & Matthews, 1974.)