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Transcription profiling by array of Legionella pneumophila ppGpp knockout and relA over expression mutants.

ABSTRACT: Effect of ppGpp on transcript level after RelA overexpression (90 min). The cells are constructed in our lab. They are knock-out mutants in the production of ppGpp(guanosine tetraphosphate) of Legionella, complemented with an over-expression vector containing the relA gene to express the RelA protein (responsible for the ppGpp production). The aim is to follow the gene response in Legionella pneumophila after induction of ppGpp at different time points.

ORGANISM(S): Legionella pneumophila

SUBMITTER: Carmen Buchrieser

PROVIDER: E-MTAB-211 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Distinct roles of ppGpp and DksA in Legionella pneumophila differentiation.

Dalebroux Zachary D ZD Yagi Brian F BF Sahr Tobias T Buchrieser Carmen C Swanson Michele S MS

Molecular microbiology 20100228 1

To transit between hosts, intracellular Legionella pneumophila transform into a motile, infectious, transmissive state. Here we exploit the pathogen's life cycle to examine how guanosine tetraphosphate (ppGpp) and DksA cooperate to govern bacterial differentiation. Transcriptional profiling revealed that during transmission alarmone accumulation increases the mRNA for flagellar and Type IV-secretion components, secreted host effectors and regulators, and decreases transcripts for translation, me ...[more]

PMID: 20199605

Similar Datasets

Transcription profiling by array of Legionella pneumophilaEfter RelA overexpression.

Project description:Effect of ppGpp on transcript level after RelA overexpression (45 min). The cells are constructed in our lab. They are knock-out mutants in the production of ppGpp(guanosine tetraphosphate) of Legionella, complemented with an over-expression vector containing the relA gene to express the RelA protein (responsible for the ppGpp production). The aim is to follow the gene response in Legionella pneumophila after induction of ppGpp at different time points.

2010-04-13 | E-MTAB-222 | biostudies-arrayexpress

Transcription profiling of Legionella pneumophila letS insertion mutants

Project description:L. pneumophila Lp02 (thyA hsdR rpsL; MB110) is a virulent thymine auxotroph derived from the Philadelphia 1 clinical isolate and was the parental strain for all constructed mutants. To construct a letS (lpg1912) insertion mutant that lacks pflaG, the letS locus containing the transposon insertion was amplified from MB417 and transferred to Lp02 by natural competence, resulting in strain MB416 (27). Within each experiment, similar culture densities were used to analyze strain phenotypes.

2010-04-01 | E-MTAB-200 | biostudies-arrayexpress

(p)ppGpp controls stringent factors by exploiting antagonistic allosteric coupling between catalytic domains

Project description:Escherichia coli RelA is a ribosomal factor with strong (p)ppGpp synthesis activity that is dramatically activated in the presence of deacylated tRNA in the ribosomal A-site. RelA is a unique enzyme in that it is directly positively regulated by its product, alarmone nucleotide (p)ppGpp. Using HDX-MS, mulecular docking, biochemsitry, and microbiology approaches, we localise the (p)ppGpp binding site of E. coli RelA and uncover the molecular mechanism of RelA regulation by the alarmone.

2021-08-11 | PXD019953 | Pride

Expression analysis of Saccharomyces cerevisiae INVSc1 under accumulation of the bacterial alarmone ppGpp

Project description:ppGpp accumulation caused by ectopic expression of RelA in Saccharomyces cerevisiae gave rise to marked changes in gene expression with both upregulation and downregulation, including changes in mitochondrial gene expression. The most prominent upregulation (38-fold) was detected in the function-unknown hypothetical gene YBR072C-A, followed by many other known stress-responsive genes. ppGpp acuumulation resulted in enhancement of tolerance against various stress stimuli, such as osmotic stress, ethanol, hydrogen peroxide, and high temperature. A two chip study using total RNA recoverd from the Saccharomyces cerevisiae TN2080 (accumulating ppGpp) and TN2077 (vector control) grown to mid-growth phase (8h) in SC-uracil medium. The plasmid pYC2/CT (V5-epitope tag vector) was used as a vector to express Sj-RSH.

2012-05-30 | E-GEOD-38239 | biostudies-arrayexpress

Transcription profiling of Streptomyces coelicolor strain M653 in a time series after ppGpp synthesis activation by thiostrepton

Project description:ppGpp synthesis was activated in exponentially growing cells of Streptomyces coelicolor in the absence of amino acid starvation by using a strain (M653 [DrelA tipAp::relA(1.46kb)]) that expresses a truncated portion of relA under the control of a thiostrepton-inducible promoter. Samples were harvested at 30 min intervals following induction for comparison to a control set of samples from aliquots of the same cultures that were not induced.

2007-08-01 | E-MEXP-1119 | biostudies-arrayexpress

Interaction-based capture and identification of protein effectors of ppGpp, the master regulator of bacterial growth

Project description:The small nucleotide ppGpp is a conserved regulatory molecule in prokaryotes that tunes growth rate in response to nutrient availability. How ppGpp regulates growth remains poorly defined. In Escherichia coli, ppGpp directly binds RNA polymerase to modulate transcription, but it likely has other targets. Here, we develop a capture-compound mass spectrometry approach to identify more than 50 putative ppGpp targets that control many key cellular processes, including translation, polyamine synthesis, and nucleotide metabolism. We validate several proteins as bona fide ppGpp targets in vivo. In particular, we demonstrate that ppGpp inhibits the enzymes Gsk and PurF to block purine salvage and de novo synthesis, respectively. Our results indicate that downregulating ATP and GTP synthesis is a critical facet of growth control by ppGpp. Collectively, our results provide new insight into ppGpp-based growth control in E. coli and candidates for future exploration. The capture compounds developed will also enable identification of ppGpp targets in other species, including many pathogens.

2018-10-04 | PXD010402 | Pride

RelA/(p)ppGpp regulates the glucose-starvation induced stringent response in the zoonostic Streptococcus suis

Project description:Growth curves and (p)ppGpp accumulation assays showed that RelA inactivation could influence S. suis growth and led to incapacity of (p)ppGpp synthesis during glucose starvation. To identify the roles of RelA/(p)ppGpp in global gene regulation in S. suis, we compared the transcriptional profiles of SC-19 [a (p)ppGpp+ strain] and ΔrelA [a (p)ppGpp0 strain during glucose starvation] in both glucose-abundant and -deficient CDM in exponential phase by microarray analysis. A less stringent cut-off limit, 2-fold change, was used. qRT-PCR validation displayed the same trends observed in the microarrays relA mutant strain and its parents strain SC-19 were cultured in both glucose-abundant CDM (CDM containing 1% glucose) and glucose-deficient CDM (CDM containing 2% glucose) respectively. The bacteriain in exponential phase were collected for microarray analysis. Three independent experiments were performed.

2015-08-08 | E-GEOD-70092 | biostudies-arrayexpress

Activation of the stringent response by loading of tRNA-RelA complexes at the ribosomal A-site

Project description:RelA/SpoT Homologs (RSHs) are ubiquitous bacterial enzymes that synthesize and hydrolyze (p)ppGpp in response to environmental challenges. Bacteria cannot survive in hosts and produce infection without activating the (p)ppGpp-mediated stringent response, but it is not yet understood how the enzymatic activities of RSHs are controlled. Using UV crosslinking and deep sequencing, we show that Escherichia coli RelA [(p)ppGpp synthetase I] interacts with uncharged tRNA during steady-state cell growth without being activated. Amino acid starvation leads to loading of cognate tRNA·RelA complexes at vacant ribosomal A-sites. In turn, RelA is activated and synthesizes (p)ppGpp. Mutation of a single, conserved residue in RelA simultaneously prevents tRNA binding, ribosome binding, and activation of RelA, showing that all three processes are interdependent. Our results support a model in which (p)ppGpp synthesis occurs by ribosome-bound RelA interacting with the Sarcin-Ricin Loop of 23S rRNA.

2018-04-05 | GSE108758 | GEO

Transcription profiling of Legionella longbeachae in exponential and postexponential phases

Project description:To know the expression pattern in exponential and postexponential phases of Legionella longbeachae growth curve

2010-02-25 | E-MTAB-199 | biostudies-arrayexpress

The Role of RelA of Streptococcus mutans in Global Control of Gene Expression

Project description:The production of (p)ppGpp by Streptococcus mutans UA159 is catalyzed by three gene products, RelA, RelP and RelQ. Here, we investigate the role of the RelA (Rel) homologue of S. mutans in the stringent response and in global control of gene expression. RelA of S. mutans was shown to synthesize pppGpp in vitro from GTP and ATP in the absence of added ribosomes, as well as in vivo in an E. coli relA-spoT mutant. Mupirocin (MUP) was shown to induce high levels of (p)ppGpp production in S. mutans in a relA-dependent manner, with a concommitant reduction in GTP pools. Keywords: (p)ppGpp, nutrient starvation, biofilm, virulence, stress

2007-11-14 | GSE9382 | GEO

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