Construction of hybrid plasmids containing the lysA gene of Escherichia coli: Studies of expression in Escherichia coli and Saccharomyces cerevisiae

Chenais, Joël; Richaud, Catherine; Ronceray, Jeanine; Cherest, Hélène; Surdin-Kerjah, Yolande; Patte, Jean‐Claude

doi:10.1007/bf00293935

Cited by 19 publications

(4 citation statements)

References 32 publications

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“…Intracellular induction of these S. flexneri genes would provide the bacterium with an increased ability to acquire phosphate and iron in the eukaryotic cytosol. Likewise, Henle cell induction of the S. flexneri lysA and bioA genes suggests that free lysine and biotin may be limiting in the eukaryotic intracellular environment, since in E. coli these genes are repressed by lysine and biotin, respectively (4,7). Although identification of individual SIIGs and the in vitro signals that regulate these genes may provide insights into Shigella physiology and the nature of the eukaryotic cytosolic environment, the in vivo situation is likely to be extremely complex, as there are numerous eukaryotic cytosolic signals and multiple bacterial systems to respond to these signals.…”

Section: Discussionmentioning

confidence: 99%

Identification of Chromosomal Shigella flexneri Genes Induced by the Eukaryotic Intracellular Environment

2002

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Upon entry into the eukaryotic cytosol, the facultative intracellular bacterium Shigella flexneri is exposed to an environment that may necessitate the expression of particular genes for it to survive and grow intracellularly. To identify genes that are induced in response to the intracellular environment, we screened a library containing fragments of the S. flexneri chromosome fused to a promoterless green fluorescent protein gene (gfp). Bacteria containing promoter fusions that had a higher level of gfp expression when S. flexneri was intracellular (in Henle cells) than when S. flexneri was extracellular (in Luria-Bertani broth) were isolated by using fluorescence-activated cell sorting. Nine different genes with increased expression in Henle cells were identified. Several genes (uhpT, bioA, and lysA) were involved in metabolic processes. The uhpT gene, which encoded a sugar phosphate transporter, was the most frequently isolated gene and was induced by glucose-6-phosphate in vitro. Two of the intracellularly induced genes (pstS and phoA) encode proteins involved in phosphate acquisition and were induced by phosphate limitation in vitro. Additionally, three iron-regulated genes (sufA, sitA, and fhuA) were identified. The sufA promoter was derepressed in iron-limiting media and was also induced by oxidative stress. To determine whether intracellularly induced genes are required for survival or growth in the intracellular environment, we constructed mutations in the S. flexneri uhpT and pstS genes by allelic exchange. The uhpT mutant could not use glucose-6-phosphate as a sole carbon source in vitro but exhibited normal plaque formation on Henle cell monolayers. The pstS mutant had no apparent growth defect in low-phosphate media in vitro but formed smaller plaques on Henle cell monolayers than the parent strain. Both mutants were as effective as the parent strain in inducing apoptosis in a macrophage cell line.

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Section: Discussionmentioning

confidence: 99%

Identification of Chromosomal Shigella flexneri Genes Induced by the Eukaryotic Intracellular Environment

2002

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“…Lysine biosynthesis, one of the important components of metabolic networks, is a pathway starting with aspartate and runs through the diaminopimelate pathway in E. coli. 6 From previous studies, [7][8][9][10][11][12][13][14][15] we have obtained an elementary understanding of the lysine biosynthesis network structure (Fig. 1).…”

Section: Introductionmentioning

confidence: 99%

“…The conversion of meso-DAP to lysine is catalyzed by DAP decarboxylase, the product of lysA. 7 The transcription of lysA is repressed by lysine. meso-DAP is synthesized from aspartic acid through the successive reactions of eight enzymes.…”

Section: Introductionmentioning

confidence: 99%

Dynamic change in promoter activation during lysine biosynthesis in Escherichia coli cells

Yamada

Nagahisa

et al. 2008

Mol. BioSyst.

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We investigated the expression dynamics of genes involved in lysine biosynthesis in Escherichia coli cells to obtain a quantitative understanding of the gene regulatory system. By constructing reporter strains expressing the green fluorescence protein (gfp) gene under the control of the promoter regions of those genes associated with lysine biosynthesis, time-dependent changes in gene expression in response to changes in lysine concentration in the medium were monitored by flow cytometry. Five promoters involved in lysine biosynthesis respond to the changes in lysine concentration in the medium. For these five promoters, time-dependent gene expression data were fitted to a simple dynamical model of gene expression to estimate the parameters of the gene regulatory system. According to the fitting parameters, dapD shows a significantly larger coefficient of repression than the other genes in the lysine synthesis pathway, which indicates the weak binding activity of the repressor to the dapD promoter region. Moreover, there is a trend that the closer an enzyme is to the start of the lysine biosynthesis pathway, the smaller its maximal promoter activity is. The results provide a better quantitative understanding of the expression dynamics in the lysine biosynthesis pathway.

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“…When trying to obtain bacterial strains that overproduce metabolites, one of the several possible approaches that can be followed is the amplification of the copy number of those genes acting as the limiting steps. The lysine pathway of Escherichia coli has been exhaustively studied in this way, showing that the method may be useful to increase the production of this amino acid (4,5,8,18,(22)(23)(24). However, the production of L-lysine by E. coli cells is much lower than that obtained with Corynebacterium and Brevibacterium strains (25).…”

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confidence: 99%

Cloning and expression in Escherichia coli of genes involved in the lysine pathway of Brevibacterium lactofermentum

Márquez¹,

Sousa²,

Sánchez³

1985

J Bacteriol

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The Brevibacterium lactofermentum genes which complement Escherichia coli lysA and asd-1 mutants were identified, respectively, as a 1.9-kilobase PstI-ClaI fragment and a 2.5-kilobase PstI fragment by cloning into pBR325. Southern blot transfers show hybridization to chromosomal fragments of identical size. The putative B. lactofermentum asd and lysA products are 44 and 48 kilodaltons, respectively.

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Construction of hybrid plasmids containing the lysA gene of Escherichia coli: Studies of expression in Escherichia coli and Saccharomyces cerevisiae

Cited by 19 publications

References 32 publications

Identification of Chromosomal Shigella flexneri Genes Induced by the Eukaryotic Intracellular Environment

Identification of Chromosomal Shigella flexneri Genes Induced by the Eukaryotic Intracellular Environment

Dynamic change in promoter activation during lysine biosynthesis in Escherichia coli cells

Cloning and expression in Escherichia coli of genes involved in the lysine pathway of Brevibacterium lactofermentum

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