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1.
J Appl Microbiol ; 130(6): 2018-2028, 2021 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-33152175

RESUMO

AIM: Erwinia amylovora is the causal agent of fire blight, a devastating disease of apples and pears. This study determines whether the E. amylovora guanine-hypoxanthine transporter (EaGhxP) is required for virulence and if it can import the E. amylovora produced toxic analogue 6-thioguanine (6TG) into cells. METHODS AND RESULTS: Characterization of EaGhxP in guanine transport deficient Escherichia coli reveals that it can transport guanine, hypoxanthine and the toxic analogues 8-azaguanine (8AG) and 6TG. Similarly, EaGhxP transports 8AG and 6TG into E. amylovora cells. EaGhxP has a high affinity for 6TG with a Ki of 3·7 µmol l-1 . An E. amylovora ⊿ghxP::Camr strain shows resistance to growth on 8AG and 6TG. Although EaGhxP is expressed during active disease propagation, it is not necessary for virulence as determined on immature apple and pear assays. CONCLUSIONS: EaGhxP is not required for virulence, but it does import 6TG into E. amylovora cells. SIGNIFICANCE AND IMPACT OF THE STUDY: As part of the disease establishment process, E. amylovora synthesizes and exports a toxic guanine derivative 6TG. Our results are counter intuitive and show that EaGhxP, an influx transporter, can move 6TG into cells raising questions regarding the role of 6TG in disease establishment.


Assuntos
Erwinia amylovora/metabolismo , Guanina/metabolismo , Hipoxantina/metabolismo , Proteínas de Transporte de Nucleobases/metabolismo , Tioguanina/metabolismo , Azaguanina/metabolismo , Erwinia amylovora/enzimologia , Erwinia amylovora/genética , Escherichia coli/genética , Escherichia coli/metabolismo , Malus/microbiologia , Proteínas de Transporte de Nucleobases/genética , Doenças das Plantas/microbiologia , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo
2.
Plant J ; 21(4): 317-27, 2000 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-10758483

RESUMO

The maize mutation high chlorophyll fluorescence 60-muTable 1 (hcf60-m1), generated through Activator (Ac) tagging, has insufficient photosynthetic electron transport. Here we show that the Hcf60 gene encodes a protein with substantial amino acid similarity to plant plastid and bacterial ribosomal small subunit protein 17 (RPS17) proteins. The lack of detectable HCF60 transcripts in mutant leaves, and insertion of the transposed Ac element 17 bp upstream of the start of translation in the mutated locus, suggest that little if any RPS17 is produced. The mutant phenotype is consistent with reduced plastid translation. Seedling lethal hcf60-m1 plants display temperature and light-dependent chlorophyll deficiencies, a depletion of plastid rRNA pools, and few high-molecular-weight polysomal complexes. Growth under moderate light conditions (27 degrees C, 100 microE m-2 sec-1) allows for substantial chlorophyll accumulation in mutant leaves, yet the number of functional photosystem II complexes appears low. Nevertheless, the presence of a limited but intact C4 system indicates that some plastid translation occurs.


Assuntos
Proteínas de Arabidopsis , Cloroplastos/genética , Genes de Plantas , Mutação , Proteínas de Plantas/genética , Proteínas Ribossômicas/genética , Zea mays/genética , Sequência de Aminoácidos , Bactérias/genética , Sequência de Bases , Cloroplastos/metabolismo , Dados de Sequência Molecular , Mutagênese Insercional , Fotossíntese , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/química , Proteínas Ribossômicas/química , Alinhamento de Sequência , Homologia de Sequência de Aminoácidos
3.
Development ; 125(15): 2813-23, 1998 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-9655804

RESUMO

The maize leafbladeless1 (lbl1) mutant displays a variety of leaf and plant phenotypes. The most extreme manifestation in the leaf is the formation of radially symmetric, abaxialized leaves due to a complete loss of adaxial cell types. Less severe phenotypes, resulting from a partial loss of adaxial cell identity, include the formation of ectopic laminae at the boundary between abaxialized, mutant sectors on the adaxial leaf surface and the bifurcation of leaves. Ectopic laminae and bifurcations arise early in leaf development and result in an altered patterning of the leaf along the proximodistal axis, or in complete duplication of the developing organ. Leaf-like lateral organs of the inflorescences and flowers show similar phenotypes. These observations suggest that Lbl1 is required for the specification of adaxial cell identity within leaves and leaf-like lateral organs. Lbl1 is also required for the lateral propagation of leaf founder cell recruitment, and plays a direct or indirect role in the downregulation of the homeobox gene, knotted1, during leaf development. Our results suggest that adaxial/abaxial asymmetry of lateral organs is specified in the shoot apical meristem, and that formation of this axis is essential for marginal, lateral growth and for the specification of points of proximodistal growth. Parallels between early patterning events during lateral organ development in plants and animals are discussed.


Assuntos
Padronização Corporal/genética , Regulação da Expressão Gênica no Desenvolvimento , Genes de Plantas , Mutação , Folhas de Planta/crescimento & desenvolvimento , Proteínas de Plantas , Zea mays/genética , Alelos , Diferenciação Celular , Genes Homeobox , Genes Recessivos , Proteínas de Homeodomínio/biossíntese , Meristema/citologia , Meristema/genética , Meristema/crescimento & desenvolvimento , Modelos Genéticos , Morfogênese/genética , Folhas de Planta/citologia , Folhas de Planta/genética , Zea mays/crescimento & desenvolvimento
4.
Plant Cell ; 8(3): 463-75, 1996 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-8721750

RESUMO

Adjacent bundle sheath and mesophyll cells cooperate for carbon fixation in the leaves of C4 plants. Mutants with compromised plastid development should reveal the degree to which this cooperation is obligatory, because one can assay whether mesophyll cells with defective bundle sheath neighbors retain C4 characteristics or revert to C3 photosynthesis. The leaf permease1-mutable1 (lpe1-m1) mutant of maize exhibits disrupted chloroplast ultrastructure, preferentially affecting bundle sheath choroplasts under lower light. Despite the disrupted ultrastructure, the metabolic cooperation of bundle sheath and mesophyll cells for C4 photosynthesis remains intact. To investigate this novel mutation, the Activator transposon-tagged allele and cDNAs corresponding to the Lpe1 mRNA from wild-type plants were cloned. The Lpe1 gene encodes a polypeptide with significant similarity to microbial pyrimidine and purine transport proteins. An analysis of revertant sectors generated by Activator excision suggests that the Lpe1 gene product is cell autonomous and can be absent up to the last cell divisions in the leaf primordium without blocking bundle sheath chloroplast development.


Assuntos
Cloroplastos/fisiologia , Genes de Plantas , Proteínas de Membrana Transportadoras/metabolismo , Proteínas de Plantas/metabolismo , Zea mays/fisiologia , Sequência de Aminoácidos , Sequência de Bases , Divisão Celular , Clonagem Molecular , Cinética , Luz , Proteínas de Membrana Transportadoras/biossíntese , Proteínas de Membrana Transportadoras/genética , Dados de Sequência Molecular , Mutagênese , Fases de Leitura Aberta , Fenótipo , Folhas de Planta , Proteínas de Plantas/biossíntese , Proteínas de Plantas/genética , Purinas/metabolismo , RNA Mensageiro/biossíntese , RNA Mensageiro/metabolismo , Homologia de Sequência de Aminoácidos , Zea mays/genética
6.
Mol Cell Biol ; 11(1): 322-8, 1991 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-1986228

RESUMO

An initiation site for meiotic gene conversion is located in the promoter region of the ARG4 locus in Saccharomyces cerevisiae. We have tested the hypothesis that the initiation site is identical with the promoter by making a series of small deletions that remove specific promoter elements. Disruption of most promoter elements does not lower the level of gene conversion in ARG4, and analysis of RNA levels at the time of recombination in meiosis reveals no direct correlation between the level of ARG4 transcript and the level of gene conversion in ARG4. However, deletion of a tract of 14 A residues located at the peak of the gene conversion gradient decreases the number of gene conversion events stimulated by the initiation site to 25 to 35% of the normal level. We conclude that the poly(dA.dT) tract is responsible for most but not all of the high levels of meiotic gene conversion observed in ARG4.


Assuntos
Poli dA-dT/genética , Regiões Promotoras Genéticas , Recombinação Genética , Saccharomyces cerevisiae/genética , Transcrição Gênica , Clonagem Molecular , Análise Mutacional de DNA , DNA Fúngico/genética , Conversão Gênica , Meiose
7.
Gene ; 96(2): 177-80, 1990 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-2176637

RESUMO

The Saccharomyces cerevisiae gene (THR1) encoding homoserine kinase (HK; EC 2.7.1.39) was cloned by complementation in yeast. Disruption of the THR1 gene results in threonine auxotrophy in yeast. Comparison of the amino acid sequences of yeast and bacterial HKs reveals substantial similarity.


Assuntos
Fosfotransferases (Aceptor do Grupo Álcool) , Fosfotransferases/genética , Saccharomyces cerevisiae/genética , Sequência de Aminoácidos , Bactérias/enzimologia , Bactérias/genética , Sequência de Bases , Genes Fúngicos , Teste de Complementação Genética , Dados de Sequência Molecular , Fosfotransferases/metabolismo , Saccharomyces cerevisiae/enzimologia , Saccharomyces cerevisiae/fisiologia , Homologia de Sequência do Ácido Nucleico , Treonina/metabolismo
8.
Genetics ; 126(4): 813-22, 1990 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-1981763

RESUMO

We have constructed eight restriction site polymorphisms in the DED81-ARG4 region and examined their behavior during meiotic recombination. Tetrad analysis reveals decreasing gradients of gene conversion on both sides of the initiation site for meiotic recombination at the ARG4 locus, extending on one side into the ARG4 gene, and on the other side into the adjacent DED81 gene. Gene conversion events can extend in both directions from the initiation site as the result of a single meiotic event. There is a second gradient of gene conversion in DED81, with high levels near the 5' end of the gene and low levels near the middle of the gene. The peaks of gene conversion activity for the DED81 and ARG4 gradients map to regions where double-strand breaks are found during meiosis. The implications of these results for models of meiotic gene conversion are discussed.


Assuntos
Conversão Gênica , Recombinação Genética , Saccharomyces cerevisiae/genética , Mapeamento Cromossômico , Cromossomos Fúngicos , Cruzamentos Genéticos , Genes Fúngicos , Marcadores Genéticos , Meiose , Polimorfismo de Fragmento de Restrição , Saccharomyces cerevisiae/citologia
9.
Proc Natl Acad Sci U S A ; 87(19): 7653-7, 1990 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-2217196

RESUMO

We have used denaturant-gel electrophoresis to provide a physical demonstration of heteroduplex DNA in the products of yeast meiosis. We examined heteroduplex formation at arg4-nsp, a G.C----C.G transversion that displays a moderately high level of postmeiotic segregation. Of the two possible arg4-nsp/ARG4 mismatches (G.G and C.C), only C.C was detected in spores from mismatch repair-competent (Pms1+) diploids. In contrast, C.C and G.G were present at nearly equal levels in spores from Pms1- diploids. These results confirm previous suggestions that postmeiotic segregation spores contain heteroduplex DNA at the site of the marker in question, that C.C is repaired less frequently than is G.G, and that the PMS1 gene product plays a role in mismatch correction. Combined with the observation that Pms1+ ARG4/arg4-nsp diploids produce 3 times more 3+:5m (wildtype:mutant) tetrads (+, +, +/m, m) than 5+:3m tetrads (+, +/m, m, m), these results indicate that, during meiosis, formation of heteroduplex DNA at ARG4 involves preferential transfer of the sense (nontranscribed) strand of the DNA duplex.


Assuntos
DNA Fúngico/genética , Ácidos Nucleicos Heteroduplexes/genética , Saccharomyces cerevisiae/genética , Sequência de Bases , Cruzamentos Genéticos , DNA Fúngico/análise , Genótipo , Meiose , Dados de Sequência Molecular , Ácidos Nucleicos Heteroduplexes/análise , Hibridização de Ácido Nucleico , Sondas de Oligonucleotídeos , Plasmídeos , Saccharomyces cerevisiae/citologia , Esporos Fúngicos/fisiologia
10.
Nature ; 338(6210): 35-9, 1989 Mar 02.
Artigo em Inglês | MEDLINE | ID: mdl-2537472

RESUMO

An initiation site for meiotic gene conversion has been identified in the promoter region of the ARG4 gene of Saccharomyces cerevisiae. The chromosome on which initiation occurs is the recipient of genetic information during gene conversion.


Assuntos
Conversão Gênica , Genes Fúngicos , Meiose , Saccharomyces cerevisiae/genética , Deleção Cromossômica , Elementos de DNA Transponíveis , Heterozigoto , Homozigoto , Regiões Promotoras Genéticas , Saccharomyces cerevisiae/citologia
11.
Nature ; 338(6210): 87-90, 1989 Mar 02.
Artigo em Inglês | MEDLINE | ID: mdl-2645528

RESUMO

It has been proposed that the initiation of meiotic recombination involves either single-strand or double-strand breaks in DNA. It is difficult to distinguish between these on the basis of genetic evidence because they give rise to similar predictions. All models invoke initiation at specific sites to explain polarity, which is a gradient in gene conversion frequency from one end of a gene to the other. In the accompanying paper we describe the localization of an initiation site for gene conversion to the promoter region of the ARG4 gene of the yeast Saccharomyces cerevisiae. Here, we show that a double-strand break appears at the ARG4 recombination initiation site at the time of recombination, and that the broken DNA molecules end in long single-stranded tails.


Assuntos
Conversão Gênica , Genes Fúngicos , Meiose , Saccharomyces cerevisiae/genética , Plasmídeos , Regiões Promotoras Genéticas , Recombinação Genética , Mapeamento por Restrição , Saccharomyces cerevisiae/citologia
12.
Cell ; 45(4): 529-36, 1986 May 23.
Artigo em Inglês | MEDLINE | ID: mdl-3518944

RESUMO

We have examined the effect of physical length on the mitotic segregation of artificial chromosomes and fragments of natural yeast chromosomes. Increasing the length of artificial chromosomes decreases the rate at which they are lost during mitosis. We have made fragments of chromosome III by integrating new telomeres at different positions along the length of the chromosome. Chromosome fragments of 42 and 72 kb behave like artificial chromosomes: they are lost in mitosis much more frequently than natural chromosomes. In contrast, a chromosome fragment of 150 kb is as mitotically stable as the full-length chromosome from which it is derived. The structural instability of a short dicentric artificial chromosome demonstrates that, although short artificial chromosomes segregate poorly in mitosis, they do attach to the mitotic spindle. We discuss these results in the context of a model in which chromosome segregation is directed by the intercatenation of the segregating DNA molecules.


Assuntos
Cromossomos/ultraestrutura , Mitose , Saccharomyces cerevisiae/fisiologia , Não Disjunção Genética , Plasmídeos , Saccharomyces cerevisiae/genética , Fuso Acromático/fisiologia
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