A messenger RNA for capsid protein isolated from tobacco mosaic virus-infected tissue

Siegel, Albert; Hari, V.; Montgomery, Ilene Nowicki; Kolacz, Kathryn

doi:10.1016/0042-6822(76)90397-4

Cited by 86 publications

(26 citation statements)

References 23 publications

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“…One might speculate then what the biological significance of the bidirectional encapsidation may be. It is known that a short piece of RNA containing the 3'-terminal 750 nucleotides of TMV RNA occurs in infected tissues and that this RNA functions as messenger of coat protein in the wheat germ cell-free systems (22,23). Lebeurier et al (10) hypothesized that this RNA may be produced by replication of the free 3' tail of the RNA of incompletely assembled virus.…”

Section: Discussionmentioning

confidence: 99%

Kinetics of biphasic reconstitution of tobacco mosaic virus in vitro.

Fukuda¹,

Ohno²,

Okada³

et al. 1978

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

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The kinetics of the in vitro reconstitution of tobacco mosaic virus from its RNA and protein were studied by measuring the increase in turbidity, the development of ribonuclease-resistant infectivity, the encapsidation of the terminal ends of the RNA, and the growth of rod length. Recently, several laboratories independently reached the conclusion that reconstitution starts at an internal region of the RNA, about 800-1000 nucleotides from the 3' terminus, and that the elongation occurs bidirectionally (7-10). In an early phase of the reaction, the rod elongation toward the 5' end is much favored over that toward the 3' end (9-11). The present study deals with kinetics of the bidirectional elongation of TMV rods during the reconstitution reaction, studied by measuring turbidity, infectivity, RNase resistance of the two ends of RNA, and rod length. The results show that reconstitution takes place in two successive phases that differ not only in the direction, but also in the rate of rod elongation. Rods with a length of 260 nm were identified as the product of the early process of reconstitution.MATERIALS AND METHODS Preparation of TMV, TMV Protein, TMV RNA, and Tritium-Labeled TMV RNA. TMV, a Japanese common strain OM, was purified by polyethylene glycol precipitation (12) followed by differential centrifugation. TMV protein and RNA were isolated by the acetic acid method (13) and by phenolbentonite extraction (14), respectively. TMV RNA radioactively labeled at its termini was prepared by periodate oxidation and subsequent reduction with tritiated sodium borohydride (8). Both the 5'-and the 3'-ribosyl ends of TMV RNA were labeled with about the same efficiency.The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U. S. C. §1734 solely to indicate this fact.Reconstitution Reaction. A protein disk preparation was obtained by incubation of the protein (6.6 mg/ml) for 24 hr in 0.1 M phosphate buffer (pH 7.0) at 20°. Reconstitution reactions were carried out in the same buffer at 200 by mixing the protein disk preparation (5.6 mg/ml) with a mixture of TMV RNA (0.13 mg/ml) and tritium-labeled TMV RNA (0.05 mg/ml). At intervals, aliquots were withdrawn to determine the rod lengths, the ribonuclease resistance of tritium-labeled termini, and the infectivity.Turbidimetric Measurement during Reconstitution Reaction. The reconstitution reaction during the initial 90 min of the reaction was followed by the increase in absorbance at 310 nm with a Gilford model 250 spectrophotometer.Rod Length Measurements by Electron Microscopy. An aliquot from the reaction mixture was diluted 1:100 with icecold 0.01% (wt/vol) bovine serum albumin solution, and minute drops of the mixture were placed on carbon-coated electron microscope grids. The grids were immediately transferred to a desiccator and kept under reduced pressure overnight. The specimens were then negatively stained with 1% (wt/vol) phosphotungst...

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Section: Discussionmentioning

confidence: 99%

Kinetics of biphasic reconstitution of tobacco mosaic virus in vitro.

Fukuda¹,

Ohno²,

Okada³

et al. 1978

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

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“…Total ssRNA from mockinoculated and MNSV-infected cucumber cotyledons was prepared as described (Siegel et al, 1976). These RNAs plus MNSV virion RNA and DNA size standards were denatured with glyoxal and electrophoresed as described (McMaster & Carmichael, 1977).…”

Section: Methodsmentioning

confidence: 99%

Nucleotide sequence and genomic organization of melon necrotic spot virus

Riviere

Rochon

1990

Journal of General Virology

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“…Having demonstrated that the seco-pregnane steroid and its glycosides can restrict the long-distance movement and, to a lesser extent, cell-to-cell spread of TMV in its host plants, we sought to investigate the molecular mechanisms underlying these inhibitory effects. TMV CP and MP are translated, respectively, from a sgRNA, originally called low molecular mass component (17,18) and from the I 2 sgRNA (19), both produced during virus replication and 3Ј-coterminal with the TMV genomic RNA. The accumulation level of the CP sgRNA or I 2 sgRNA is thus a key determinant for the amount of CP or MP synthesized.…”

Section: Seco-pregnane Steroid and Glycosides Selectivelymentioning

confidence: 99%

Seco -pregnane steroids target the subgenomic RNA of alphavirus-like RNA viruses

Wang²,

et al. 2007

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

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Plants have evolved multiple mechanisms to selectively suppress pathogens by production of secondary metabolites with antimicrobial activities. Therefore, direct selections for antiviral compounds from plants can be used to identify new agents with potent antiviral activity but not toxic to hosts. Here, we provide evidence that a class of compounds, seco-pregnane steroid glaucogenin C and its monosugar-glycoside cynatratoside A of Strobilanthes cusia and three new pantasugar-glycosides of glaucogenin C of Cynanchum paniculatum, are effective and selective inhibitors to alphavirus-like positive-strand RNA viruses including plant-infecting tobacco mosaic virus (TMV) and animal-infecting Sindbis virus (SINV), eastern equine encephalitis virus, and Getah virus, but not to other RNA or DNA viruses, yet they were not toxic to host cells. In vivo administration of the compounds protected BALB/c mice from lethal SINV infection without adverse effects on the mice. Using TMV and SINV as models, studies on the action mechanism revealed that the compounds predominantly suppress the expression of viral subgenomic RNA(s) without affecting the accumulation of viral genomic RNA. Our work suggested that the viral subgenomic RNA could be a new target for the discovery of antiviral drugs, and that seco-pregnane steroid and its four glycosides found in the two medicinal herbs have the potential for further development as antiviral agents against alphavirus-like positive-strand RNA viruses.antiviral drugs ͉ Strobilanthes cusia ͉ Cynanchum atratum V iruses replicate in host cells by hijacking the host-cell metabolic machinery. It is therefore difficult for antiviral therapies to inhibit only viral functions but spare host cellular processes. Nevertheless, great progress has been achieved over the past decades in antiviral research and therapy to reduce morbidity and mortality in virus-infected individuals (1, 2), and many antiviral drugs have been approved in recent years, most of which targeted either viral proteins or host cellular proteins. However, some of these compounds might induce the resistance of viruses to them and cause cell toxicity (3-5). The continuing emergence of highly pathogenic viruses like avian influenza viruses and severe acute respiratory syndrome coronavirus underscores the importance of advancing the search for effective antiviral agents. Plants have evolved constitutive and inducible defense mechanisms by producing a vast array of secondary metabolites against various microbial pathogens (6). Many herbaceous plants have been used in traditional Chinese medicine to treat viral diseases (7). It is conceivable that antiviral compounds would occur in plants as part of their innate defense arsenal, and the vast assortment of secondary metabolites could serve as a large pool for screening for previously undescribed antiviral compounds with a selective target spectrum and nontoxic to the host plants. Here, we provide evidence that a class of compounds, the seco-pregnane steroid and its glycosides from Strobilanthe...

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A messenger RNA for capsid protein isolated from tobacco mosaic virus-infected tissue

Cited by 86 publications

References 23 publications

Kinetics of biphasic reconstitution of tobacco mosaic virus in vitro.

Kinetics of biphasic reconstitution of tobacco mosaic virus in vitro.

Nucleotide sequence and genomic organization of melon necrotic spot virus

Seco -pregnane steroids target the subgenomic RNA of alphavirus-like RNA viruses

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