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At least 865 records · Page 48Linked to original sources
PubMed1994

Complete nucleotide sequence and genome organization of a tobamovirus infecting cruciferae plants.

Genomic RNA sequence of a tobamovirus infecting cruciferae plants (cr-TMV) was determined. The RNA is composed of 6312 nucleotides and contains four ORFs encoding the proteins of 122K (ORF1), 178K (ORF2), 29K (ORF3) and 18K (capsid protein, ORF4). ORF4 overlaps ORF3 by 74 nucleotides and the overlapping region can be folded into a stable hairpin structure. The 3'-terminal region of the cr-TMV RNA preceding the tRNA-like structure was shown to form six potentially stable pseudoknots.

Amino Acid Sequence↗
PubMed1982

Localization of antigenic determinants of a viral protein by inhibition of enzyme-linked immunosorbent assay (ELISA) with tryptic peptides.

A new method is described for localizing antigenic determinants in globular proteins by means of inhibition of enzyme-linked immunosorbent assay (ELISA) with tryptic peptides. The inhibitory activity of 10 tryptic peptides of the coat protein of 2 strains of tobacco mosaic virus (TMV) was demonstrated by inhibition of ELISA and verified by inhibition of complement fixation tests. These results bring to 7 the number of antigenic determinants of TMV protein that have been defined.

Amino Acid Sequence↗
PubMed1987

Correlation of co-ordinated amino acid substitutions with function in viruses related to tobacco mosaic virus.

Sequence data are available for the coat proteins of seven tobamoviruses, with homologies ranging from at least 26% to 82%, and atomic co-ordinates are known for tobacco mosaic virus (TMV) vulgare. A significant spatial relationship has been found between groups of residues with identical amino acid substitution patterns. This strongly suggest that their location is linked to a particular function, at least in viruses identical with the wild-type for these residues. The most conserved feature of TMV is the RNA binding region. Core residues are conserved in all viruses or show mutations complementary in volume. The specificity of inter-subunit contacts is achieved in different ways in the three more distantly related viruses.

Amino Acid Sequence↗
PubMed1989

Visualization of alpha-helices in tobacco mosaic virus by cryo-electron microscopy.

We have used tobacco mosaic virus (TMV) as a test specimen, in order to develop techniques for the analysis of high-resolution structural detail in electron micrographs of biological assemblies with helical symmetry. It has previously been shown that internal details of protein structure can be visualized by processing electron micrographs of unstained specimens of extended two-dimensional crystalline arrays. However, the techniques should in principle be applicable to other periodic specimens, such as assemblies with helical symmetry. We show here that data to spacings better than 10 A can be retrieved from electron images of frozen hydrated TMV. The three-dimensional computed map agrees well with that derived from X-ray diffraction and shows the two pairs of alpha-helices forming the core of the coat subunit, the C alpha-helix and the viral RNA. The results demonstrate that it is possible to determine detailed internal structure in helical particles.

Freezing↗
PubMed1989

Visualization of protein-nucleic acid interactions in a virus. Refined structure of intact tobacco mosaic virus at 2.9 A resolution by X-ray fiber diffraction.

The structure of tobacco mosaic virus (TMV) has been determined by fiber diffraction methods at 2.9 A resolution, and refined by restrained least-squares to an R-factor of 0.096. Protein-nucleic acid interactions are clearly visible. The final model contains all of the non-hydrogen atoms of the RNA and the protein, 71 water molecules, and two calcium-binding sites. Viral disassembly is driven by electrostatic repulsions between the charges in two carboxyl-carboxylate pairs and a phosphate-carboxylate pair. The phosphate-carboxylate pair and at least one of the carboxyl-carboxylate pairs appear to be calcium-binding sites. Nucleotide specificity, enabling TMV to recognize its own RNA by a repeating pattern of guanine residues, is provided by two guanine-specific hydrogen bonds in one of the three base-binding sites.

Amino Acids↗
PubMed1991

The signal for a leaky UAG stop codon in several plant viruses includes the two downstream codons.

Expression of the RNA replicase domain of tobacco mosaic virus (TMV) and certain protein-coding regions in other plant viruses, is mediated by translational readthrough of a leaky UAG stop codon. It has been proposed that normal tobacco tyrosine tRNAs are able to read the UAG codon of TMV by non-conventional base-pairing but recent findings that stop codons can also be bypassed as a result of extended translocational shifts (tRNA hopping) have encouraged a re-examination. In light of the alternatives, we investigated the sequences flanking the leaky UAG codon using an in vivo assay in which bypass of the stop codon is coupled to the transient expression of beta-glucuronidase (GUS) reporter genes in tobacco protoplasts. Analysis of GUS constructions in which codons flanking the stop were altered allowed definition of the minimal sequence required for read through as UAG-CAA-UUA. The effects of all possible single-base mutations in the codons flanking the stop indicated that 3' contexts of the form CAR-YYA confer leakiness and that the 3' context permits read through of UAA and UGA stop codons as well as UAG. Our studies demonstrate a major role for the 3' context in the read through process and do not support a model in which teh UAG is bypassed exclusively as a result of anticodon-codon interactions. No evidence for tRNA hopping was obtained. The 3' context apparently represents a unique sequence element that affects translation termination.

Base Composition↗
PubMed1992

Structure of the U2 strain of tobacco mosaic virus refined at 3.5 A resolution using X-ray fiber diffraction.

The structure of the U2 strain of tobacco mosaic virus (TMV) has been determined by fiber diffraction methods at 3.5 A resolution, and refined by a combination of restrained least-squares and molecular dynamics methods to an R-factor of 0.096. The structure is extremely similar to that of the common strain of TMV, with the largest differences being in the protein loop that makes up the inner surface of the virus, and in the C-terminal region on the outer surface. Differences in the inner loop can be correlated with differences in the properties of the two viruses.

Capsid↗
PubMed1985

Antiviral activity of natural and recombinant human leukocyte interferons in tobacco protoplasts.

Several purified species of human leukocyte interferon, including recombinant interferons, inhibit the multiplication of tobacco mosaic virus (TMV) in tobacco protoplasts derived from various cultivars. Viral RNA accumulation was determined by dot-blot hybridization to specific cDNA probes, and virus antigen was determined serologically. Interferon apparently inhibited both TMV-RNA replication and its expression into coat protein. However, these effects were of limited duration. Maximum effect was obtained when interferon was applied to the cells either prior to inoculation or within the first hour after inoculation. Antibodies to interferon abolished its antiviral activity in protoplasts. Tobacco protoplasts were about 1000 times more responsive to interferon than the reference animal viral-cell system and showed an "antiviral state" at a ratio of 1 molecule of interferon per cell.

DNA Replication↗
PubMed1988

Tobacco mosaic virus particles contain ubiquitinated coat protein subunits.

Virions of tobacco mosaic virus (TMV) are composed of a single strand of RNA, encapsidated in about 2130 copies of a coat protein of MW 17,500. Asselin and Zaitlin [Virology 91, 173-181 (1978)] demonstrated that virion preparations also contained small amounts of a second protein of MW 26,500, which they termed "H protein." H protein, detectable to an average frequency of one per virion, was thought to be a protein of host origin. Subsequent studies [Collmer, Vogt, and Zaitlin, Virology 126, 429-448 (1983)] showed the H protein was comprised of a backbone of TMV coat protein, linked by a postulated isopeptide bond to a small protein that probably was of host origin. The host-derived moiety of H protein is shown here to be ubiquitin, most probably coupled to the coat protein at lysine 53. This finding is based on microsequencing of the H protein, and is substantiated by immunoblotting analysis with antibodies to human ubiquitin. Conjugated ubiquitin was detected in virions of all five strains of the virus tested. To our knowledge, this is the first report of a ubiquitinated viral structural protein.

Capsid↗
PubMed1990

Replication, stability, and gene expression of tobacco mosaic virus mutants with a second 30K ORF.

A series of tobacco mosaic virus (TMV)-hybrids containing a second 30K open reading frame (ORF) inserted into different positions of the genome 3' region were constructed. These insertional mutants were used to evaluate the effects of a modified viral genome organization on replication and gene expression. They were evaluated for stability upon systemic infection and subsequent host passage using RNase protection assays. A mutant with the second 30K ORF fused in frame to two-thirds of the coat protein reading frame replicated as a free-RNA virus and produced increased amounts of the hybrid protein compared to the wild-type 30K protein, but substantially reduced amounts compared to the wild-type coat protein. A mutant with the second 30K ORF inserted between the native 30K and coat protein ORFs produced reduced amounts of 30K protein but replicated efficiently and was maintained for weeks of systemic infection before the population gradually shifted to progeny wild-type TMV. Mutants with the second 30K ORF fused behind different lengths of the coat protein subgenomic RNA promoter/leader region and inserted between the coat protein gene and the 3' nontranslated sequences replicated poorly and the mutations were not maintained during continued replication in plants.

Base Sequence↗
PubMed1991

Tobacco rattle virus RNA-1 29K gene product potentiates viral movement and also affects symptom induction in tobacco.

In order to investigate the function of the 29K protein of tobacco rattle virus (TRV), we introduced different mutations in the 29K protein gene and analyzed the biological properties of the subsequent transcripts in tobacco plants. Although none of the mutant RNAs was able to accumulate to a detectable level, the defects in the 29K protein could be complemented by coinoculation with wild-type TRV or tobacco mosaic virus (TMV). Complementation was also achieved in transgenic plants expressing the homologous TMV 30K protein which is involved in cell-to-cell movement, but without inducing distinctive symptoms. Transcripts of chimeric TRV clones containing duplicate genes for the 29K protein initiated infections with formation of necrotic lesions and the progeny retained only one copy of the gene. These experiments demonstrate that the 29K protein is not required for viral RNA replication and, because the TRV transcripts do not encode the coat protein, that the 29K and 30K proteins act on nonencapsidated RNA. In addition to potentiating viral movement, the TRV 29K protein may also play a role in symptom induction on tobacco.

Blotting, Northern↗
PubMed1992

Effects of terminal deletion mutations on function of the movement protein of tobacco mosaic virus.

A series of carboxy- and amino-terminal deletion mutations in the movement protein (MP) gene of tobacco mosaic virus (TMV) were ligated into a cloned TMV cDNA deleted for the endogenous MP gene. RNA transcripts were produced in vitro from clones carrying the various mutated MP genes. The effect of the deletion mutations on local and systemic movements of the infection was evaluated. Deletion of 9 or 33 amino acids from the carboxy terminus of the movement protein did not effect cell-to-cell movement as reflected by local lesion formation on Nicotiana tabacum cv. Xanthi NN plants. Deletion of 55 amino acids resulted in impaired MP that supported the formation of local lesions of 1 mm in diameter compared to lesions of 3-5 mm caused by the wild-type MP. Deletion of 74 amino acids (or more) from the carboxy terminus resulted in a protein that could not support virus movement. Modified viruses that contained repeated sequences in the 3' region of the MP gene lost the repeated sequences during replication and reverted to the wild type. This was evidenced by the size of the MP produced and by sequence analysis of reverse-transcribed PCR-amplified products, following infection by the modified virus. MP deleted for as few as 3 amino acids at the amino terminus could not support virus movement thus indicating that the amino-terminal domain is critical for MP activity.

Cell Wall↗
PubMed1985

Immunochemical studies of tobacco mosaic virus--VI. Attempts to localize viral epitopes with monoclonal antibodies.

The specificity of 18 monoclonal antibodies directed to tobacco mosaic virus (TMV) was studied by measuring their ability to bind to viral mutants, to other tobamoviruses, to dissociated viral subunits and to peptide fragments of the viral coat protein. The apparent binding specificity of the antibodies was dependent on the type of enzyme-linked immunosorbent assay used, probably because the antigens were disrupted or denatured when attached to the plastic surface of microtiter wells. The capacity of different monoclonal antibodies to detect single substitutions in the viral coat protein was used to delineate some of the topographic epitopes of TMV. By means of computer-generated images of the surface residues of the viral subunit, it was possible to identify certain clusters of residues involved in binding to some of the monoclonal antibodies. The results clearly illustrate the operational limitations encountered when monoclonal antibodies are used for elucidating the antigenic structure of proteins.

Amino Acid Sequence↗
PubMed1983

A simple procedure for the extraction of double-stranded RNA from virus-infected plants.

A simple procedure for the isolation of double-stranded (ds) RNA from virus-infected plants is described. The method is based on grinding plant tissue in 4% p-aminosalicylic acid and recovery of ds RNA by phenol extraction and precipitation with 30% ethanol. The presence of both negative and positive virus RNA strands in RNA fractionated in agarose gels was verified by Northern blot hybridization with polynucleotide kinase labelled genomic RNA or complementary DNA (cDNA) probes. The procedure enabled detection of three major ds RNA species (MWs 4.2, 1.05 and 0.48 X 10(6)) and at least 4 minor bands with estimated MWs of 3.5, 2.5, 2.2 and 2.0 X 10(6) in Nicotiana tabacum plants systemically infected with tobacco mosaic virus (TMV). Cucumber mosaic virus (CMV)-infected Pachystachys coccinea plants contained 2 minor bands of MWs 0.49 and 0.35 X 10(6) in addition to the previously described 4 major ds RNAs and ds CARNA 5 (MW 0.22 X 10(6)). The patterns of ds RNA are useful for diagnosing natural infections of CMV and TMV in N. glauca plants and of citrus tristeza virus in Citrus spp.

Mosaic Viruses↗
PubMed1993

Transfection of whole plants from wounds inoculated with Agrobacterium tumefaciens containing cDNA of tobacco mosaic virus.

We engineered cDNA of tobacco mosaic tobamovirus (TMV) into Agrobacterium tumefaciens for inoculation of plant cells. The resulting bacterial strains were used to transfect tobacco (Nicotiana tabacum cv. Xanthi and Xanthi/nc) with wild type and a defective virus. Lesion formation on Xanthi/nc tobacco was used to measure the timing and efficiency of transfection. Infections mediated by Agrobacterium produced lesions an average of two days later than infections produced by inoculation with virions. The addition of approximately 80 bp of non-viral sequences to the 5'-end of TMV transcripts abolished transfection. Transcripts with non-viral sequences at the 3'-end initiated infections, while precise transcript termination with a synthetic ribozyme sequence increased transfection frequencies two-fold. Culture conditions reported to induce genes of the vir region of the Agrobacterium Ti plasmid also increased the transfection frequency approximately two-fold. Therefore, in addition to the pararetroviruses and geminiviruses previously described, 'agroinoculation' may be used to infect plants with plus-sense RNA viruses.

Agrobacterium tumefaciens↗
PubMed1985

Comparison of the effects of chlorite-oxidized oxyamylose and polyacrylic acid on the multiplication of phytopathogenic viruses.

Polyacrylic acid (PAA) and chlorite-oxidized oxyamylose (COAM) inhibit the multiplication of tobacco mosaic virus (TMV) in leaf disks by up to 50%. The reduction in TMV content is time-dependent and decreases with longer time intervals between the virus infection and the application of substances. The multiplication of potato virus X (PVX) in leaf disks is not affected by either PAA or COAM. In intact plants PAA produces a strong antiviral effect on both PVX and red clover mottle virus (RCMV). The effect produced by COAM is much less pronounced, although this substance is less toxic and could be used in a higher concentration than PAA. Neither of these compounds has a significant influence on the development of virus-induced necroses in Nicotiana glutinosa, Gomphrena globosa or Phaseolus vulgaris plants when administered one day before or after virus infection.

Acrylic Resins↗
PubMed1995

Plant virus expressing hybrid coat protein with added murine epitope elicits autoantibody response.

A modified tobacco mosaic virus (TMV) was used to direct the synthesis of TMV coat protein hybrids containing a 13 amino acid sequence of the murine zona pellucida ZP3 protein. This hybrid protein was found to be robust and accumulate to high concentrations in inoculated plants. Virus-like particles containing the hybrid coat protein could be readily purified from infected plant tissue. Parenteral immunization with these virus-like particles as antigen resulted in serum antibody recognizing ZP3 epitope in C57BL/6J and BALB/cBy mice. The antibodies recognized synthetic ZP3 peptide and the authentic murine ZP3 glycoprotein. Antibodies were recruited in vivo to the zone pellucida in female mice.

Animals↗
PubMed1991

Translational efficiency and competitive ability of mRNAs with 5'-untranslated alpha beta-leader of potato virus X RNA.

The 5'-untranslated leader sequence of potato virus X (PVX) RNA (63 nucleotides apart from cap-structure) consists of two sub-sequences referred to as alpha-sequence (41 nucleotides with no G) and beta-sequence (42 nucleotides upstream from the first AUG). Computer-based folding predictions suggest that the 5'-proximal region of alpha beta-leader is unstructured. The second structural feature of alpha beta-leader is the presence of the sequences apparently complementary to the 3'-terminal region of 18S rRNA. The alpha beta-leader has been shown to strongly enhance the translation of the contiguous foreign gene (NPT1) transcripts in cell-free translation systems from rabbit reticulocytes (RRL), wheat germ (WG) and Krebs-2 ascite cell extract (KA). In competitive translation PVX, RNA strongly inhibited tobacco mosaic virus (TMV) RNA, in RRL and WG systems. No competition occurred between PVX and TMV RNAs in KA system. There was no correlation between the translational efficiency and competitive ability of PVX RNA in different cell-free translation systems. The competitive ability did not solely depend on the presence of alpha beta-leader in mRNA. We present evidence to suggest that alpha beta-leader together with about 150 bases of the coding sequence is responsible for the translation competitive ability of PVX RNA.

Base Sequence↗