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1.
Gene ; 155(2): 305-6, 1995 Apr 03.
Artigo em Inglês | MEDLINE | ID: mdl-7721109

RESUMO

The complete nucleotide sequence (8031 bp) of the DNA of cauliflower mosaic virus (CaMV) strain B29 is reported. This strain is unusual, since it infects both cruciferous and solanaceous plants. So far, from data of sequence comparisons between B29 and other CaMV strains there is no evidence for any obvious correlation between host range and distinct sequence features.


Assuntos
Vírus do Mosaico/genética , Plantas/virologia , Sequência de Aminoácidos , Sequência de Bases , Dados de Sequência Molecular , Alinhamento de Sequência
2.
Virology ; 202(2): 1043-5, 1994 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-8030207

RESUMO

The gene III product (P15) of cauliflower mosaic virus (CaMV) is a DNA binding protein in which the DNA binding activity is located on its C-terminal part. In previous work, a C-terminal processed form of P15 (P11) was detected in purified viral particles as a minor component. The full-length P15 was shown to be present and to be matured, possibly by a cysteine proteinase, in CaMV replication complexes isolated from infected turnip leaves. In this paper, we have shown that a virion-enriched fraction obtained from such replication complexes by size exclusion chromatography contained most of the P15 in its uncleaved form and was enriched in the activity responsible for its proteolysis. This enabled us to characterize better the proteinase activity (temperature and pH optimum; effect of specific inhibitors) responsible for P15 cleavage and to confirm that it corresponds to a cysteine proteinase. Based upon these observations, a purification procedure for CaMV particles was devised which impaired the cleavage of P15 into P11 and allowed the isolation of virions containing almost exclusively the noncleaved form. This finding supports our hypothesis that the CaMV gene III product could be involved in the folding of the viral genome during encapsidation.


Assuntos
Caulimovirus/metabolismo , Proteínas de Ligação a DNA/metabolismo , Proteínas Virais/metabolismo , Capsídeo/metabolismo , Caulimovirus/ultraestrutura , Endopeptidases/metabolismo , Genes Virais , Fases de Leitura Aberta , Proteínas Estruturais Virais/genética , Vírion/ultraestrutura , Replicação Viral
3.
Virology ; 197(1): 283-92, 1993 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-8212564

RESUMO

Cauliflower mosaic virus (CaMV) aphid transmission factor (ATF), produced in a baculovirus expression system, forms paracrystalline structures, as demonstrated by electron microscopic observations. Similar paracrystals were also found in CaMV-infected plants, using immunogold techniques, thus providing the first evidence of such a structure for the CaMV ATF (P18). We demonstrated that the paracrystals can be solubilized to provide an active form of the CaMV ATF which can also be reverted into the paracrystalline aggregated form. This suggests that the paracrystalline structures might act as a source of active CaMV ATF or be the form in which it is stored within the infected cells. A point mutation within the CaMV gene II (which encodes the ATF) leads to the loss of both the paracrystalline structures and the ATF activity. Hence, the paracrystalline structure seems to be a feature of the native (unmodified ) CaMV ATF.


Assuntos
Afídeos/microbiologia , Caulimovirus/metabolismo , Proteínas Virais/biossíntese , Proteínas Virais/metabolismo , Animais , Baculoviridae/genética , Brassica , Caulimovirus/genética , Cristalização , Genes Virais , Insetos , Microscopia Eletrônica , Peso Molecular , Proteínas Recombinantes/análise , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/ultraestrutura , Recombinação Genética , Mapeamento por Restrição , Transfecção , Proteínas Virais/análise
4.
Proc Natl Acad Sci U S A ; 90(4): 1470-3, 1993 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-8434006

RESUMO

We cloned in Escherichia coli truncated versions of the protein p15 encoded by open reading frame III of cauliflower mosaic virus. We then compared the ability of the wild-type p15 (129 amino acids) and the deleted p15 to bind viral double-stranded DNA genome. Deletions of > 11 amino acids in the C-terminal proline-rich region resulted in loss of DNA binding activity of wild-type p15. Moreover, a point mutation of the proline at position 118 sharply reduced the interaction between the viral protein and DNA. These results suggest that cauliflower mosaic virus p15 belongs to the family of DNA binding proteins having a proline-rich motif involved in interaction with double-stranded DNA.


Assuntos
Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Vírus do Mosaico/genética , Fases de Leitura Aberta , Proteínas Virais/genética , Proteínas Virais/metabolismo , Sequência de Aminoácidos , Sítios de Ligação , Brassica , Códon/genética , DNA Viral/genética , Escherichia coli/genética , Genoma Viral , Dados de Sequência Molecular , Mutagênese Sítio-Dirigida , Plasmídeos , Prolina , Proteínas Recombinantes de Fusão/metabolismo , Deleção de Sequência , Homologia de Sequência de Aminoácidos , beta-Galactosidase/genética , beta-Galactosidase/metabolismo
5.
Biochimie ; 75(8): 645-9, 1993.
Artigo em Inglês | MEDLINE | ID: mdl-8286436

RESUMO

The electrophoretic forms of purified cauliflower mosaic virus (CaMV), strain Cabb-S, were examined by electrophoresis on agarose gels. Three populations of viral particles were identified: a faster migrating component (the form F) and two slower migrating components (the forms S and S'). When the different forms of virions, after excision from gels, were subjected to analysis in SDS-polyacrylamide gel, the fast component consisted of the 37 and 42 kDa coat proteins whereas the slow components contained mainly the 39 kDa coat protein. However, there was no difference among the nucleic acids associated within the three forms. The biological significance of the different components is discussed.


Assuntos
Caulimovirus/química , Vírion/química , Southern Blotting , Capsídeo/isolamento & purificação , Caulimovirus/genética , DNA Viral/análise , Eletroforese em Gel de Ágar , Vírion/isolamento & purificação
6.
Res Virol ; 143(5): 361-70, 1992.
Artigo em Inglês | MEDLINE | ID: mdl-1480825

RESUMO

The major capsid protein of the cauliflower mosaic virus (CaMV) is processed in vivo. The viral aspartic proteinase that catalyses this maturation has been characterized previously and is coded by the CaMV gene V. This virus has a second capsid protein, a minor component, encoded by gene III. This protein, P3, is also processed at its C-terminus in vivo. To determine whether P3 is matured by the CaMV proteinase P5, we expressed, in Saccharomyces cerevisiae, P3, P5 and a fusion protein P7-P4, containing potential sites of cleavage. P5 was found to be involved in maturation of P7-P4 but did not cleave P3. The latter result was confirmed by experiments carried out with an in vitro translation system (the reticulocyte lysate) and with preparations of replication complexes purified from infected plants. Moreover, [N-(L-3-trans-carboxyoxiran-2-carbonyl)-L-leu cyl]-amido(4-guanido)butane, a specific inhibitor of cysteine proteinases, inhibited the maturation of P3, suggesting that the two CaMV capsid proteins are not processed by the same proteolytic event.


Assuntos
Capsídeo/metabolismo , Cisteína Endopeptidases/metabolismo , Vírus do Mosaico/metabolismo , Processamento de Proteína Pós-Traducional , Capsídeo/genética , Clonagem Molecular , Inibidores de Cisteína Proteinase/farmacologia , Genes Virais , Leucina/análogos & derivados , Leucina/farmacologia , Vírus do Mosaico/genética , Biossíntese de Proteínas , Precursores de Proteínas/metabolismo , Saccharomyces cerevisiae/genética
7.
FEBS Lett ; 287(1-2): 1-4, 1991 Aug 05.
Artigo em Inglês | MEDLINE | ID: mdl-1715279

RESUMO

Reverse transcription is not solely a retroviral mechanism. Hepadnaviruses and caulimoviruses have RNA intermediates that are reverse transcribed into DNA. Moreover non-viral retroelements, retrotransposons, use reverse transcription in their transposition. All these retroelements encode reverse transcriptase but each group developed their own expression modes capable of assuring a specific and efficient replication of their genomes.


Assuntos
RNA Viral/metabolismo , DNA Polimerase Dirigida por RNA/metabolismo , Vírus/enzimologia , Capsídeo/genética , Capsídeo/metabolismo , Elementos de DNA Transponíveis , HIV-1/enzimologia , Hepadnaviridae/enzimologia , Hepadnaviridae/genética , Retroviridae/enzimologia , Retroviridae/genética , Vírus/genética
8.
Res Virol ; 142(4): 297-302, 1991.
Artigo em Inglês | MEDLINE | ID: mdl-1796216

RESUMO

Cauliflower mosaic virus (CaMV) gene I encodes a 40-kDa protein, P1, which is thought to be involved in the cell-to-cell movement of the virus. In order to investigate its functioning, P1 was expressed in Saccharomyces cerevisiae transformed by an expression vector containing CaMV gene I. When produced in yeast, PI was 40 kDa in size and not N-glycosylated.


Assuntos
Genes Virais/genética , Vírus do Mosaico/genética , Plantas Comestíveis/microbiologia , Proteínas Virais/genética , Western Blotting , Clonagem Molecular , Expressão Gênica/genética , Vetores Genéticos/genética , Glicosilação , Saccharomyces cerevisiae/genética
9.
Virology ; 180(2): 837-41, 1991 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-1703375

RESUMO

The capsid protein and the reverse transcriptase of cauliflower mosaic virus (CaMV) are encoded by two genes (ORF IV and ORF V) that lie in different translation reading frames. A comparison can be drawn between the synthesis of both CaMV proteins and the fusion protein in a yeast retrotransposon, Ty, resulting from a +1 frameshifting event which fuses two out-of-phase ORFs encoding the structural protein and the reverse transcriptase of Ty. For this reason, we constructed a yeast expression vector containing CaMV ORF VII fused to CaMV ORF III by a fragment of 452 bp including the overlapping region of ORF IV and ORF V, ORF VII and ORF III being used as reporter genes. We characterized two proteins (22 and 50 kDa) synthesized from this plasmid in the yeast expression system. We demonstrated that the 50-kDa polypeptide is not synthesized from a +1 frameshifting event but is probably a dimeric form of the 22-kDa protein. From this result we conclude that the CaMV reverse transcriptase is not produced by a mechanism of ribosomal frameshifting.


Assuntos
Capsídeo/genética , Mutação da Fase de Leitura , Genes Virais , Vírus do Mosaico/genética , DNA Polimerase Dirigida por RNA/genética , Brassica , Elementos de DNA Transponíveis , Immunoblotting , Vírus do Mosaico/enzimologia , Fases de Leitura Aberta , Plasmídeos , Mapeamento por Restrição , Saccharomyces cerevisiae/genética
10.
J Virol ; 64(6): 2594-8, 1990 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-2186173

RESUMO

Antiserum was prepared against a synthetic peptide corresponding to the N-terminal 20 amino acids of the protein encoded by cauliflower mosaic virus (CaMV) open reading frame VII (ORF VII). This antiserum was used to detect the expression of CaMV ORF VII either in Saccharomyces cerevisiae transformed by an expression vector containing CaMV ORF VII or in CaMV-infected plants. Only in S. cerevisiae has a 14-kilodalton protein been detected.


Assuntos
Brassica/microbiologia , Genes Virais , Vírus do Mosaico/genética , Saccharomyces cerevisiae/genética , Brassica/genética , Mapeamento Cromossômico , Eletroforese em Gel de Poliacrilamida , Escherichia coli/genética , Expressão Gênica , Immunoblotting , Plasmídeos , Proteínas Virais/genética , Proteínas Virais/isolamento & purificação
11.
Virology ; 174(2): 622-4, 1990 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-2305555

RESUMO

The interaction of the gene III product, P15, of cauliflower mosaic virus with different double-stranded DNA fragments of the viral genome was investigated. The results suggest that gene III product which showed DNA binding activity is a structural protein of the viral particle.


Assuntos
DNA Viral/metabolismo , Proteínas de Ligação a DNA/análise , Genes Virais , Vírus de Plantas/genética , Proteínas Virais/análise , Sequência de Aminoácidos , Vírus de Plantas/análise , Proteínas Estruturais Virais/análise
13.
Virus Genes ; 2(4): 313-22, 1989 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-2815594

RESUMO

The cauliflower mosaic virus (CaMV) particle-associated protein kinase (PK) was shown to be a 37 kD protein in activity gels. In vitro experimental data concerning virus dephosphorylation or hyperphosphorylation suggested a possible regulation mechanism of this PK. The origin of the enzyme, either virus-encoded or from a host cell, is discussed.


Assuntos
Núcleo Celular/enzimologia , Vírus do Mosaico/enzimologia , Proteínas Quinases/metabolismo , Brassica , Eletroforese em Gel de Poliacrilamida , Peso Molecular , Vírus do Mosaico/genética , Fosforilação , Plantas/enzimologia , Proteínas Quinases/genética , Proteínas Quinases/isolamento & purificação
14.
Ann Inst Pasteur Virol ; 139(3): 263-76, 1988.
Artigo em Inglês | MEDLINE | ID: mdl-3061414

RESUMO

A CaMV DNA fragment corresponding to nucleotides 2200-3992 and including the coding sequence (2200-3670) of open reading frame IV was inserted in the pTG908 prokaryotic expression vector. In the recombinant pTG-IV plasmid, ORF IV, which codes for the coat protein precursor, was fused to the N-terminal coding sequence of the lambda CII gene, which is under transcriptional control of the lambda PL promoter. The expected fusion protein CII-ORF IV had a calculated molecular weight of 58.4 Kd. Nevertheless, temperature induction of the PL promoter resulted in synthesis of a major 76-Kd fusion protein: the coat protein precursor migrated abnormally on SDS polyacrylamide gel.


Assuntos
Genes Virais , Vírus do Mosaico/genética , Proteínas Virais/genética , Western Blotting , Clonagem Molecular , DNA Recombinante , Escherichia coli/genética , Vetores Genéticos , Peso Molecular , Precursores de Proteínas/genética
15.
Virology ; 163(2): 503-8, 1988 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-3354204

RESUMO

Gene I product of cauliflower mosaic virus was immunodetected in a cell-wall-enriched fraction from infected turnip leaves in addition to its detection in viroplasms and replication complexes. The immunoreaction was carried out with an antiserum raised against a 15 amino acid long synthetic peptide corresponding to the carboxy-terminus of potential gene I protein (P1). The presence of P1 in different subcellular fractions was investigated as a function of time during viral multiplication. At late infection times, P1 was found only in the cell-wall-enriched fraction.


Assuntos
Parede Celular/análise , Genes Virais , Vírus do Mosaico/genética , Plantas/microbiologia , Proteínas Virais/análise , Vírus do Mosaico/fisiologia , Fragmentos de Peptídeos/síntese química , Fragmentos de Peptídeos/imunologia , Frações Subcelulares/análise , Proteínas Virais/fisiologia , Replicação Viral
16.
Plant Mol Biol ; 11(3): 271-6, 1988 May.
Artigo em Inglês | MEDLINE | ID: mdl-24272340

RESUMO

Antisera against the N-terminal and C-terminal parts of the potential ORF IV product were used to analyse extracts from CaMV-infected turnip leaves by immunoblotting. Polypeptides of 87, 83, 82, 60 and 57 kDa were detected. The origin of these proteins is discussed.

17.
EMBO J ; 5(10): 2433-8, 1986 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-16453711

RESUMO

Using electron microscopic immunocytochemical staining we demonstrate that the product of gene III of cauliflower mosaic virus (CaMV) is associated with viral particles. Furthermore, a fusion protein, expressed in bacteria, consisting of the N-terminus of beta-galactosidase and the complete gene III protein of CaMV showed DNA-binding activity. From these two results, we discuss the possible function of this viral polypeptide.

18.
J Gen Virol ; 67 ( Pt 7): 1491-8, 1986 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-3723112

RESUMO

Mutants of cauliflower mosaic virus (CaMV), generated in vitro by modification of recombinant DNA plasmids containing the viral genome, either retained the ability to induce disease symptoms on turnip plants, produced less severe symptoms or failed to induce symptoms. Wild-type symptoms were produced by a variant CaMV DNA of the Cabbage S isolate that had 4 bp in open reading frame (ORF) III replaced with a 16 bp sequence. Less severe symptoms, due to a delay in symptom appearance relative to inoculation with wild-type DNA, were induced by a mutant with a frameshift mutation in ORF II (pSA103). CaMV DNA, recovered from plants infected with pSA103, contained a second mutation which restored the original translation reading frame. Nucleic acid hybridization to 'squishes' of leaf tissue from plants that had been inoculated with mutant DNAs that included DNAs modified in each of the six major ORFs of CaMV DNA revealed that only those plants that appeared diseased had detectable CaMV nucleic acid in uninoculated leaves. Replicated CaMV DNA was also not detected in non-encapsidated and virion DNA fractions from inoculated leaves of non-diseased plants.


Assuntos
Vírus do Mosaico/patogenicidade , Sequência de Bases , Replicação do DNA , DNA Viral/genética , Vírus do Mosaico/genética , Vírus do Mosaico/fisiologia , Hibridização de Ácido Nucleico , Virulência , Replicação Viral
19.
Gene ; 48(2-3): 277-86, 1986.
Artigo em Inglês | MEDLINE | ID: mdl-3557131

RESUMO

Pairs of heterologous cauliflower mosaic virus (CaMV) genomes cloned in pBR322, one having a defective genome and both restricted at the same pBR322 cloning site, generate recombinant molecules in infected cells when co-inoculated on plants. Analysis of the restriction pattern of the isolated recombinant CaMV DNAs indicated that the intergenomic recombination may be explained by dimerization of two heterologous CaMV molecules and transcription into a hybrid 35S RNA responsible for replication of the recombinant genomes.


Assuntos
DNA Viral/genética , Vírus do Mosaico/genética , Recombinação Genética , Sequência de Bases , Replicação do DNA , DNA Recombinante/fisiologia , DNA Viral/fisiologia , Modelos Genéticos , Vírus do Mosaico/fisiologia , Conformação de Ácido Nucleico , Replicação Viral
20.
Plant Mol Biol ; 5(5): 281-9, 1985 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-24306919

RESUMO

A class of mutants of cauliflower mosaic virus (CaMV) DNA was distinguished based on its members' ability to induce symptoms when coinoculated on plants with other CaMV DNAs mutant at a different locus. Three mutants, one each in open reading frame I, III, and VI had this ability. A second class of mutant DNAs did not induce symptoms unless combined with a mutant DNA of the first class. Viral DNA extracted from diseased plants was shown by restriction enzyme digestion to have lost the mutant alleles. When turnip plants were inoculated with a recombining mutant derived from DNA of the Cabbage S isolate and a mutant derived from DNA of a different isolate, a heterogeneity in the viral DNA extracted from the diseased plants was detected by restriction enzyme analysis. Restriction analysis of cloned representatives of this heterogeneous population revealed regions consistent with repair of heteroduplexes formed during general recombination between duplex DNAs. Some regions consistent with this mechanism or with recombination by strandswitching during reverse transcription were found.

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