Evidence that tobacco mosaic virus particles disassemble contranslationally in vivo

Shaw, J. G.; Plaskitt, Kitty A.; Wilson, T. M. A.

doi:10.1016/0042-6822(86)90329-6

Cited by 82 publications

(36 citation statements)

References 16 publications

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“…Such effects have changed the tightly bound helical assembly into a loosely bound "beads on a string" structure that is held together by encircling (but not as strongly binding to) the viral RNA, making possible the cotranslational stripping of CP by ribosomes during disassembly (6,(26)(27)(28).…”

Section: Resultsmentioning

confidence: 99%

“…The lower calcium concentration and the higher pH in the plant cell relative to the extracellular environment allow the first several subunits of CP to fall off so that ribosome-mediated "cotranslational uncoating" can occur, due to the low affinity between CP and 5′ RNA in which guanines are few (6,(26)(27)(28). However, assembly and disassembly of TMV take place in the same plant cell; the same environmental conditions must accommodate the two seemingly opposite processes.…”

Section: Discussionmentioning

confidence: 99%

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Hydrogen-bonding networks and RNA bases revealed by cryo electron microscopy suggest a triggering mechanism for calcium switches

Zhou

2011

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

114

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Helical assemblies such as filamentous viruses, flagella, and F-actin represent an important category of structures in biology. As the first discovered virus, tobacco mosaic virus (TMV) was at the center of virus research. Previously, the structure of TMV was solved at atomic detail by X-ray fiber diffraction but only for its dormant or high-calcium-concentration state, not its low-calcium-concentration state, which is relevant to viral assembly and disassembly inside host cells. Here we report a helical reconstruction of TMV in its calcium-free, metastable assembling state at 3.3 Å resolution by cryo electron microscopy, revealing both protein side chains and RNA bases. An atomic model was built de novo showing marked differences from the high-calcium, dormant-state structure. Although it could be argued that there might be inaccuracies in the latter structure derived from X-ray fiber diffraction, these differences can be interpreted as conformational changes effected by calcium-driven switches, a common regulatory mechanism in plant viruses. Our comparisons of the structures of the low-and highcalcium states indicate that hydrogen bonds formed by Asp116 and Arg92 in the place of the calcium ion of the dormant (highcalcium) state might trigger allosteric changes in the RNA basebinding pockets of the coat protein. In turn, the coat protein-RNA interactions in our structure favor an adenine-X-guanine (A*G) motif over the G*A motif of the dormant state, thus offering an explanation underlying viral assembly initiation by an AAG motif.H elical assemblies represent a very important class of biological structures, performing various tasks for the proper functions of life or executing malicious functions in disease. For example, protein helical assemblies such as cytoskeletal networks and muscle fibers are essential components in our bodies (1, 2). Other helical assemblies include bacterial flagella and secretion systems. Helical viruses, such as tobacco mosaic virus (TMV), account for a major fraction of the virus kingdom. Such helical structures, bearing a special kind of 2D periodicity and potential flexibility, are difficult for structural determination to atomic resolution by conventional methods including X-ray crystallography and NMR. Thus, the molecular mechanisms of action in many of these systems are not understood to atomic detail. Even for TMV, the first virus ever isolated and extensively studied (3) by both X-ray diffraction (e.g., 4-8) and cryo electron microscopy (cryoEM) (9, 10), we still do not know its structure at atomic detail in a state that is relevant to its assembly and disassembly processes, and the underlying mechanisms of these processes remain unknown.The viral RNA of TMV is infectious by itself, but RNA on its own is very unstable. The addition of a 17-kDa coat protein (CP) around the RNA protects the RNA from degrading and makes TMV very stable. The assembly of a rod-shaped virion begins at a two-turn helical CP complex, called the 20S aggregate (11), which recognizes the initiation moti...

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Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Hydrogen-bonding networks and RNA bases revealed by cryo electron microscopy suggest a triggering mechanism for calcium switches

Zhou

2011

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

114

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“…Because viral genomes must form single-stranded templates that are largely free of coat protein at some stage during the infection process, depurination of virus-related nucleic acids could provide direct protection against infection. The presence of high concentrations of RIPs within the cell wall (30,36,59) could facilitate RIP-viral nucleic acid contact; alternatively, RIPs could enter the cytoplasm together with the virions via mechanical damage to the cell wall and plasma membrane and act on viral RNA during cotranslational disassembly (62,63). Because viroids lack the protein capsid that protects almost all conventional viruses, one might expect them to be particularly sensitive to RIPs.…”

Section: Discussionmentioning

confidence: 99%

The N-Glycosidase Activity of the Ribosome-inactivating Protein ME1 Targets Single-stranded Regions of Nucleic Acids Independent of Sequence or Structural Motifs

Park¹,

Vepachedu²,

Owens

et al. 2004

Journal of Biological Chemistry

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ME 1 , a type I ribosome-inactivating protein (RIP), belongs to a family of enzymes long believed to possess rRNA N-glycosidase activity directed solely at the universally conserved residue A4324 in the sarcin/ricin loop of large eukaryotic and prokaryotic rRNAs. We have investigated the effect of modifying the structure of nonribosomal RNA substrates on their interaction with ME 1 and other RIPs. ME 1 was shown to depurinate a variety of partially denatured nucleic acids, randomly removing adenine residues from single-stranded regions and, to a lesser extent, guanine residues from wobble basepairs in hairpin stems. A defined sequence motif was not required for recognition of non-paired adenosines and cleavage of the N-glycosidic bond. Substrate recognition and ME 1 activity appeared to depend on the physical availability of nucleotides, and denaturation of nucleic acid substrates increased their interaction with ME 1 . Pretreatment of mRNA at 75°C rather than 60°C, for example, lowered the apparent K D from 87.1 to 73.9 nM, making it more vulnerable to depurination by RIPs. Exposure to ME 1 in vitro completely abolished the infectivity of partially denatured RNA transcripts of the potato spindle tuber viroid, suggesting that RIPs may target invading nucleic acids before they reach host ribosomes in vivo. Our data suggest that the extensive folding of many potential substrates interferes with their ability to interact with RIPs, thereby blocking their inactivation by ME 1

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“…This might indicate that, after vector transmission of BYV, exposure of the particle to the environment of a damaged phloem cell triggers dissociation of the p24 tail, thus making the 5h end of the RNA accessible for the ribosomes. Possibly, the main part of the genome would still be coated by the p22 at this stage, and expression of the 5h-terminal gene(s) should proceed along with the stripping of the particle by the translating ribosomes, in accord with the co-translational disassembly model proposed for tobacco mosaic virus (Wilson, 1984 ;Shaw et al, 1986). Such stepwise ' unwrapping ' of the closterovirus particles might serve to protect their RNA genomes, which otherwise would be easy targets for the cell nucleases.…”

mentioning

confidence: 90%

The minor coat protein of beet yellows closterovirus encapsidates the 5' terminus of RNA in virions.

Zinovkin¹,

Wilhelm²,

Agranovsky³

1999

Journal of General Virology

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Filamentous particles of beet yellows closterovirus (BYV) are built of two related capsid proteins, of which the minor species, p24, forms a 75 nm tail at one end of the virion. In the present work, we used polyclonal antibodies against p24 for isolating the ' tailed ' virion segments from sonicated BYV particle preparations. The [γ-32 P]ATP-labelled RNA obtained from the antibody-selected particle segments consistently showed stronger hybridization with the 5h-terminal BYV cDNA clones than with the 3h-terminal cDNA clones. These data clearly indicate that it is the 5h-terminal portion of the closterovirus RNA genome that is encapsidated by p24.Elongated plant viruses, although variable in their particle length, flexibility and stability, all have a single RNA molecule in a helical arrangement encapsidated by many identical coat protein (CP) subunits encoded by a single viral gene (Matthews, 1991). Recently, a striking deviation from this basic structure has been described : the flexuous filamentous particles of beet yellows closterovirus (BYV) were found to consist of two related CPs arranged in a ' rattlesnake ' structure (Agranovsky et al., 1995). The particles of citrus tristeza closterovirus (CTV) proved to have a similar morphologically polar structure (Febres et al., 1996). The positive-strand RNA genomes of BYV and CTV encompass tandem arrays of the genes for the major and minor CPs (p22 and p24 in BYV, and p25 and p27 in CTV, respectively) that are likely to have resulted from gene duplication (Boyko et al., 1992 ; Pappu et al., 1994). The CP duplicate genes have been identified in all other mono-and bipartite closterovirus genomes sequenced so far, and their presence is considered a hallmark of the group (Klaassen et al., 1995 ;Keim-Konrad & Jelkmann, 1996 ;Zhu et al., 1998 ; reviewed in Dolja et al., 1994 ;Agranovsky, 1996).Using immunoelectron microscopy with antibodies specific to full-sized BYV p24 or a unique N-terminal peptide thereof, Author for correspondence : Alexey Agranovsky.Fax j7 95 939 31 81. e-mail aaa!closter.genebee.msu.su the protein has been found to form a 75 nm tail at one end of the 1370 nm BYV particles ; comparison of these lengths gives a rough value of 0n85 kb for the p24-coated portion of the 15n5 kb BYV genomic RNA (Agranovsky et al., 1995). Consistent with this, the length of the p27 tails on the 2000 nm particles of CTV was determined to be 75-85 nm (Febres et al., 1996). It has been conjectured that this ' rattlesnake ' structure is shaped by a mechanism utilizing an internal origin of assembly in the closterovirus RNA, and that the p24 tail may be involved in specific interactions with receptors in the aphid vector and\or the host plant cells (Agranovsky, 1996). As a first step towards understanding the closterovirus assembly mechanism, we sought to determine the BYV genomic RNA extremity coated by the p24. To this end, the BYV particles were broken into small pieces which were segregated with p22-and p24-specific antibodies. RNA fragments obtained from these antibody-...

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Evidence that tobacco mosaic virus particles disassemble contranslationally in vivo

Cited by 82 publications

References 16 publications

Hydrogen-bonding networks and RNA bases revealed by cryo electron microscopy suggest a triggering mechanism for calcium switches

Hydrogen-bonding networks and RNA bases revealed by cryo electron microscopy suggest a triggering mechanism for calcium switches

The N-Glycosidase Activity of the Ribosome-inactivating Protein ME1 Targets Single-stranded Regions of Nucleic Acids Independent of Sequence or Structural Motifs

The minor coat protein of beet yellows closterovirus encapsidates the 5' terminus of RNA in virions.

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