Search PubMed⌕ Search

Biomedical subjects

G Lebeurier

Publications and source records attributed to G Lebeurier.

At least 37 records · Page 2Linked to original sources
PubMed1984

Expression of a putative plant viral gene in Escherichia coli.

A recombinant plasmid, pCB300, was constructed which carries a cauliflower mosaic virus (CaMV) DNA insert corresponding to nucleotides 1825-2280, including the coding sequence (1830-2219) of open reading frame III (ORF III). This CaMV DNA insert was fused with the amino-terminal portion of the beta-galactosidase gene. Transcription of the hybrid gene is controlled by the lac promoter, which is repressed in Escherichia coli strain JM103 and can be induced by isopropylthio-beta-D-galactoside (IPTG). When the promoter is derepressed, cells harboring the chimeric plasmid produce an Mr 16 000 fusion protein. This protein is immunodetected by antibodies raised against an amino terminal synthetic peptide of 19 amino acids corresponding to a sequence predicted from the nucleotide sequence of ORF III.

Cloning, Molecular↗
PubMed1984

Detection in vivo of a new gene product (gene III) of cauliflower mosaic virus.

Cauliflower mosaic virus DNA contains six major open reading frames (ORFs). As only the mRNA corresponding to the transcription of gene VI and its translation product have been isolated, the identification in infected plants of products corresponding to the five other putative genes remains to be established. The present paper reports the detection of an ORF III product by means of antibodies raised against an NH(2)-terminal synthetic peptide of 19 amino acids corresponding to a sequence predicted from the nucleotide sequence of ORF III. The detection of this gene product raises the question of the mechanism of its expression.

Journal Article↗
PubMed1983

Electron microscopic studies of the different topological forms of the cauliflower mosaic virus DNA: knotted encapsidated DNA and nuclear minichromosome.

Cauliflower mosaic virus (CaMV) DNA exists under different topological forms in infected plants. First, the population of encapsidated CaMV DNA molecules appears heterogeneous when analysed by gel electrophoresis. The electron microscopic study reported here reveals that CaMV virion DNA contains simple and multiple topological knots. Second, a supercoiled DNA form never found in virions exists as a chromatin-like nucleoprotein complex with nucleosome subunits in the nuclei of infected leaves. The compaction ratio of the minichromosomes is compatible with the nucleosomal structure, the number of nucleosomes (41.0 +/- 2.5) is in keeping with the length of the viral genome.

Cell Nucleus↗
PubMed1982

Identification by immunoprecipitation of cauliflower mosaic virus in vitro major translation product with a specific serum against viroplasm protein.

A highly specific antiserum was prepared against purified cauliflower mosaic virus viroplasm-protein (VmP). A virus specific in vitro major translation product (TPmaj), encoded by the 19S poly(A) RNA fraction from cauliflower mosaic virus infected turnip leaves, was recognized by this antiserum. The N-terminal sequence of TPmaj corresponds to the sequence following the first in-phase initiation codon in gene VI of the cauliflower mosaic virus genome. Both VmP and TPmaj have blocked termini and probably start from the same AUG codon.

Journal Article↗
PubMed1982

In vivo recombination of cauliflower mosaic virus DNA.

LIGATION AND RECOMBINATION OF THE DNA OF CAULIFLOWER MOSAIC VIRUS (CAMV) IS DEMONSTRATED BY THE FOLLOWING EXPERIMENTS: (i) Ligation: Different noninfectious fragments of the CaMV genome (obtained after insertion into plasmid pBR322 followed by enzymatic excision) regained infectivity when mixtures of them were used to inoculate their host. The symptom appearance was delayed by comparison with a typical CaMV infection, and only the newly formed leaves were affected. (ii) Recombination: Pairs of noninfectious recombinant full-length CaMV genomes (integrated into pBR322 at different restriction endonuclease sites) regained infectivity upon simultaneous inoculation of a sensitive host. The symptomatology of the resulting infection was indistinguishable from that of a typical CaMV infection. We show that progeny DNA had the same characteristics (size, structure, restriction endonuclease digestion pattern) as bona fide CaMV DNA, and that the vector pBR322 had been completely eliminated. A cloned tandem dimer of CaMV DNA with a partial deletion similarly was infectious in the plant assays. This system should be useful to study the expression of mutant genomes, thus allowing characterization of the CaMV genes.

Journal Article↗
PubMed1980

Infectivities of native and cloned DNA of cauliflower mosaic virus.

Infectivity assays on turnips reveal that (i) cauliflower mosaic virus (CaMV) DNA, whether circular or linear, is as infectious as the complete virus; (ii) linear DNA obtained with restriction enzymes from the native CaMV DNA has the same specific infectivity as when first cloned in plasmid (pBR322) or bacteriophage (lambda gtWES) vectors and then restricted at the cloning site; (iii) in all cases studied mosaic symptoms are accompanied by virus production. DNA isolated from these viruses is again circular and possesses the three "gaps" characteristic of CaMV DNA. The cloned CaMV DNA, when linked to the vector DNA, is noninfectious or exhibits very low infectivity.

Cloning, Molecular↗
PubMed1980

Restriction map of native and cloned cauliflower mosaic virus DNA.

Cloned CaMV DNA replicates faithfully in Escherichia coli, since the restriction map of the cloned DNA can be superimposed over that of the native viral DNA. However, some short fragments were difficult to detect in the restricted native viral DNA, whereas they formed clear bands when derived from cauliflower mosaic virus (CaMV) DNA clones propagated in the E. coli host. Apparently, the small fragments that carry variable-length single-stranded gaps present only in native viral DNA, give rise to diffuse weak bands difficult to recognize in gels. Comparison of maps for several CaMV strains permits evaluation of their possible evolutionary relationship.

Chromosome Mapping↗
PubMed1979

Polar uncoating of tobacco mosaic virus (TMV) with dimethylsulfoxide (DMSO) and subsequent reassembly of partially stripped TMV.

Increasing concentrations of dimethylsulfoxide (DMSO) strip tobacco mosaic virus (TMV) stepwise from the 3'end. The RNA tail increases in length up to 2,000 nucleotides (nu) reaching a region of very strong protein-RNA affinity. Thereafter, uncoating occurs from the other end and produces a second RNA tail 500 nu long. Further stripping of TMV proceeds from both ends, the long tail increasing in length up to 4,000 nu and the short one increasing more moderately and remaining below 2,000 nu. The region of strongest protein-RNA affinity is located between 4,000 and 5,000 nu away from the 3' end. Using the same conditions as for in vitro TMV reassembly, it is possible to recoat the RNA tails with viral protein preferentially in the 5' direction. The advantages of DMSO in studies of TMV protein-RNA interactions are discussed.

Dimethyl Sulfoxide↗
PubMed1978

Physical map of DNA from a new cauliflower mosaic virus strain.

The restriction enzymes AluI, BamHI, BglII, EcoRI, HindIII, and SalI have been used to characterize and map a new cauliflower mosaic virus strain (Cabb-S). These fragments have been ordered by examining their overlapping regions after double enzymatic digestion. The single SalI cleavage site was chosen as the point of origin. We compare this strain with those already described.

Base Sequence↗
PubMed1977

Inside-out model for self-assembly of tobacco mosaic virus.

Incompletely reconstituted particles of tobacco mosaic virus treated with moderate concentrations of dimethylsulfoxide prior to electron microscopy have two visible tails of unencapsidated RNA, a long one and a short one. The short tail has a constant length of 720 +/- 80 nucleotides in incomplete particles of diverse size and probably corresponds to the 3'OH end of the RNA. The length of the long 5' tail is inversely related to the length of the unfinished rod. Surprisingly, both tails appear to protrude from the same end of the particle. These observations suggest that the strand of RNA coming out of one end of the particle loops back along the length of the rod so as to appear at the opposite end. The "looped-back" tail almost certainly passes down the empty central channel of the incomplete rod. The findings are compatible with a model of tobacco mosaic virus assembly in which the RNA inserts itself beneath layers of incoming protein from within the central channel.

Binding Sites↗
PubMed1975

Location of the cistron of the tobacco mosaic virus coat protein.

Treatment of tobacco mosaic virus (TMV) RNA with T1 RNase under mild conditions cuts the RNA molecule into a large number of fragments, only a few of which may be specifically recognized by disks of TMV protein. It has been shown elsewhere that these specifically recognized RNA fragments are a part of the coat protein cistron, the portion coding for amino acids 95 to 129 of the coat protein. It is reported that different size classes of partially uncoated virus particles were prepared by limited reconstitution between TMV RNA and protein or by partial stripping of intact virus with DMSO. Both procedures produce nucleoprotein rods in which the 5'-terminal portion of the RNA is encapsidated and the 3'-terminal region is free. The free and the encapsidated portions of the RNA were each tested for the ability to give rise to the aforesaid specifically recognized fragments of the coat protein cistron upon partial T1 RNase digestion. It was found that only the 3'-terminal third of the virus particle need to be uncoated in order to expose the portion of the RNA molecule from which these fragments are derived. We conclude, therefore, that the coat protein cistron is situated upon the 3'-terminal third of the RNA chain, i.e. within 2000 nucleotides of the 3'-end.

Genes↗
PubMed1975

The uncoating of native and reconstituted TMV by dimethylsulphoxide: the polarity of stripping.

The effect of increasing concentrations of dimethylsulphoxide (DMSO) on the stability of native and reconstituted tobacco mosaic virus (TMV) has been investigated by means of electron microscopy and infectivity assay. The removal of protein subunits from TMV by DMSO was found to be a stepwise process. Subunits are first removed from the 3'OH end of the rod but at about 72% DMSO the site at which uncoating occurs shifts to the 5' end. Studies with reconstituted and partially reconstituted TMV confirm the stepwise nature of the uncoating process. Complete uncoating of reconstituted TMV was found to occur at smaller DMSO concentrations than that of native TMV.

Dimethyl Sulfoxide↗