Response of Escherichia coli to acetate in presence of 100 mM methyl α-D-glucopyranoside
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ABSTRACT:
ORGANISM(S): Escherichia coli K-12
SUBMITTER: Brice Enjalbert
PROVIDER: E-MTAB-11717 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
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Project description:E. coli MG1655 was grown on M9 minimal medium supplemented with 15mM glucose and either 0, 10, 50 or 100 mM acetate. Transcriptomic analysis from exponential growing cells was performed using Agilent microarray A-MEXP-2232. This work was done with the help of students during practicals. Consequently, it is of special interest since we have four biological replicates done over four years and two technical replicates per biological replicate (n=8 per condition). We therefore thank all the INSA-Toulouse students that participated in this project: Leidy Caraballo, Xavier Caron, Pauline Chanut, Sarah Guiziou, Ngoc Thu Hang Pham, Diane Barbay, Céline Ben Hassen, Thomas Cerutti, Cécile Roland, Sarah Srour, Audrey Baylet, Mathilde Beraud, Claudie Bosc, Lilas Courtot, Violaine Dolfo, Anna Kaci, Manon Chevallot-Beroux, Sarah Colom, Fanny Leclerc, Zihan Liao, Grégoire Quinet and Mélina Vaurs.
2020-08-21 | E-MTAB-9086 | biostudies-arrayexpress
Project description:Dihydroxyacetone (DHA) is an attractive molecule produced in a wide range of industries . DHA is found among all the kingdoms as an intermediate of various metabolic pathways and can be used as a carbon source by many organisms. The bacterium Escherichia coli is able to grow on DHA as the sole carbon source albeit at a low growth rate (0.1 h-1). If the topology of DHA metabolic network has been characterized, the function and regulation of the metabolic pathways involved in DHA metabolism remain unsolved. Here, we aimed to better understand DHA metabolism in E. coli BW25113 by exploring its transcriptional pattern on DHA versus glucose. We also studied the pattern of three DHA pathway mutants (dhaKLM, ptsA and glpK).
2019-06-03 | E-MTAB-7666 | biostudies-arrayexpress
Project description:In this project we used the strain E. coli BW25113 ΔfrmA::frt (Δ(araD-araB)567, ΔlacZ4787(::rrnB-3), λ-, rph-1, Δ(rhaD-rhaB)568, hsdR514, ΔfrmA(::frt)) from the Keio collection (Baba et al. 2006). The plasmid {pSEVA424_mdh-das} (Low-copy-number, lacIq/Ptrc promoter, RK2 ori, SmR) from Silva-Rocha et al. 2013 was integrated in the strain to express mdh (Methanol dehydrogenase) and das (Dihydroxyacetone synthase) respectively from Acinetobacter gerneri and Pichia angusta. In this experiment, we inoculated 50 mL of M9 minimal medium with 15 mM xylose and with or without 0.15 M methanol, in baffled flasks at 30°C, 150 rpm. At D600=1 and D600=2, 12mL of each culture were centrifuged 1’30 at 14000 rpm before discarding the supernatant and freezing the pellets for 15 sec in liquid nitrogen. RNA extraction were carried out using a RNeasy Midi Kit (Qiagen, Venlo, Netherlands) according to the manufacturer’s protocol and the transcriptomic experiments were run at the GeT-Biopuces Platform (Toulouse, France). Prior to the experiment, the quality of the RNA extracted was analyzed by Nanodrop and Cell Bio Analyser (Agilent Technologies, California, USA). The labelling with Cy3, hybridation on the Agilent chip and wash were carried out accordingly to the Agilent protocol and kits. We used the scanner NimbleGen MS 200 (Roche, Bâle, Suisse) with the following parameters: autogain, 5micros resolution, 532nm scan. The Agilent Feature Extraction software was used to extract the data.
2020-07-10 | E-MTAB-8909 | biostudies-arrayexpress
Project description:Because nitrogen (N) nutrition is a key determinant of plant growth, we explored the role of N availability in grafted grapevine development. Vitis vinifera cv. Cabernet Sauvignon was grafted on two rootstock genotypes known to confer high (1103 Paulsen, 1103P) and low (Riparia Gloire de Montpellier, RGM) vigour. One-year-old plants were cultivated in sand-filled pots in a greenhouse and irrigated with the control nutrient solution for 15 days of acclimation (1.6 mM N). At the end of the acclimation period (0 days post treatment (dpt)), the plants were divided in two groups of 5 plants per combination and irrigated with nutrient solutions varying only in their nitrate concentration (0.8 mM (Nitrate -) and 2.45 mM (Nitrate +)). Roots were harvested at 15 and 60 dpt. Gene expression profiling was done using the Nimblegen whole genome array with 3 biological replicates per condition to analyze the combined effect of N treatment and rootstock genotype on gene expression.
2016-04-01 | E-MTAB-3062 | biostudies-arrayexpress
Project description:This experiment studied the effect of FPP accumulation on E. coli. E. coli cells transformed with pMBIS (the S. cerevisiae mevalonate pathway enzymes converting mevalonate to FPP) and fed mevalonate produce large amounts of FPP, which causes toxicity when it accumulates. When coupled with an active amorphadiene synthase (pADS) the cells produce amorphadiene, a non-toxic isoprenoid. To accumulate FPP, but maintain similar protein burden, an amorphadiene synthase with 3 mutations to render it inactive was used (pADSmut) to accumulate FPP. E. coli was transformed with pMBIS and pADS or pMBIS and pADSMut and grown in M9+glucose with varying magnesium concentrations and fed 20 mM mevalonate and induced with 0.5 mM IPTG, then sampled at subsequent time points. This comparison is between E. coli DH1 cells accumulating FPP via the heterologous mevalonate pathway (pMBIS/pADSmut) to cells producing amorphadiene via the same pathway (pMBIS/pADS). Samples were collected 6, 10, 14, and 26.5 hr after addition of IPTG and mevalonate. One biological replicate was used. Total RNA was extracted, reverse transcribed, labeled, and hybridized to multiple slides for technical replicates.
2011-07-07 | E-GEOD-30403 | biostudies-arrayexpress
Project description:Verotoxigenic Escherichia coli (VTEC) are a leading cause of food-borne illness. Fruit and vegetables are recognised as an important source of the pathogen and can account for ~ 25 % of food-borne VTEC outbreaks, globally. The ability of VTEC to colonise leaves and roots of leafy vegetables, spinach (Spinacia oleracea) and lettuce (Lactuca sativa), was compared. The highest levels of colonisation occurred in the roots and rhizosphere, whereas colonisation of the leaves was lower and significantly different between the species. Colonisation of the leaves of prickly lettuce (L. serriola), a wild relative of domesticated lettuce, was especially poor. Differential VTEC gene expression in spinach extracts was markedly different for three tissue types, with little overlap. Comparison of expression in the same tissue type, cell wall polysaccharides, for lettuce and spinach also showed substantial differences, again with virtually no overlap. The transcriptional response was largely dependent on temperatures that are relevant to plant growth, not warm-blooded animals. The data show that VTEC adaptation to plant hosts and subsequent colonisation potential is underpinned by wholescale changes in gene expression that are specific to both plant tissue type and to the species.
2016-08-01 | E-MTAB-3249 | biostudies-arrayexpress
Project description:Transcriptomic analysis was applied to acidogenic, solventogenic and alcohologenic steady-state C. acetobutylicum cells for understanding the regulation of the metabolism of this organism. Used microarray is Clostridium acetobutylicum ATCC824 transcriptional 44k v4, correspondance to GPL10908 Phosphate-limited continuous cultures of C. acetobutylicum were performed under acidogenesis (pH 6.3, 995 mM of carbon from glucose), solventogenesis (pH 4.4, 995 mM of carbon from glucose) and alcohologenesis (pH 6.3, 498 mM of carbon from glucose and 498 mM of carbon from glycerol). Samples were collected for three biological replicates from one control strain and three metabolic mutants, respectively. Total RNA of 36 samples was extracted using an RNeasy Midi kit (Qiagen, Courtaboeuf, France) following the manufacturer's instructions with the supplementation of DNase treatment using RNase-Free DNase Set (Qiagen). Agilent microarray for Clostridium acetobutylicum ATCC 824 (4x44K AMADID 024247) was used. The RNAs were labeled with a Low Input Quick Amp Labeling kit and hybridized following a one-color microarray-based gene expression analysis protocol.
2015-07-01 | E-GEOD-69973 | biostudies-arrayexpress
Project description:Low energy states delay aging in multiple species, yet mechanisms coordinating energetics and longevity across tissues remain poorly defined. The conserved energy sensor AMP-activated protein kinase (AMPK) and its corresponding phosphatase calcineurin modulate longevity via the ‘CREB regulated transcriptional coactivator (CRTC)-1 in C. elegans. We show that CRTC-1 specifically uncouples AMPK/calcineurin mediated effects on lifespan from pleiotropic side effects by reprogramming mitochondrial and metabolic function. Strikingly, this pro-longevity metabolic state is regulated cell-nonautonomously by CRTC-1 in the nervous system. CRTC-1/CREB act antagonistically with the functional PPARα ortholog, NHR-49 to promote distinct peripheral metabolic programs. Neuronal CRTC-1 drives mitochondrial fragmentation in distal tissues and suppresses the effect of AMPK on systemic mitochondrial metabolism and longevity via a cell-nonautonomous catecholamine signal. These results demonstrate that transcriptional control of neuronal signals can override enzymatic regulation of metabolism in peripheral tissues. Central perception of energetic state therefore represents a target to promote healthy aging. Experiment was performed with three biological replicates. Gravid adults grown at 20¡C on 100 mm NG plates seeded with OP50-1 E. coli were collected and treated with hypochlorite to release eggs. Eggs were incubated overnight in M9 media to obtain L1 synchronized populations. One thousand L1 larvae were grown on a 100 mm NG plate seeded with OP50-1 E. coli. Worms were harvested for RNA extraction when L4 larval stage was reached. Animals were collected and washed extensively with M9 media to remove bacteria. Worms were then snap frozen in liquid nitrogen. RNA was extracted by five freeze/thaw cycles in Qiazol then purified by RNeasy mini kit (Qiagen). RNA quality was checked using an Agilent Technologies 2100 Bioanalyzer. All samples had an RNA integrity number of 10. cDNA libraries were prepared from 4 ugs of total RNA using the TruSeq RNA Sample Preparation v2 kit (Illumina). 50-cycle paired-end sequencing was performed on an Illumina HiSeq 2000 by the Harvard Biopolymer Core. Read quality was evaluated with FASTQC. Adapter sequences and poor quality bases (<20) were trimmed and filtered with CUTADAPT, resulting in a median of 44 million reads per replicate. These were aligned to the C. elegans genome (ce6, WS238) using TopHat version 2.0.8 (Kim et al., 2013), with a median 35 million reads mapped in proper pairs. The number of reads mapping to each gene was counted with htseq-count. Genes with less than 1 Count Per Million Reads (CPM) were discarded from further analysis. Counts were normalized for sequencing depth and RNA composition across all samples with edgeR (Robinson et al., 2010). Genes were tested for differential expression between each mutant strain and wild-type using edgeR’s glm method. For each comparison, genes with less than 5 CPM were filtered and those with at least 50% change and False Discovery Rate (FDR) of 1% or less were considered differentially expressed.
2015-02-27 | E-GEOD-58931 | biostudies-arrayexpress
Project description:A synthetic pathway for (D)-xylose assimilation was stoichiometrically evaluated regarding its potential to produce selected value-added compounds and implemented in Escherichia coli strains. The pathway proceeds via the isomerization of (D)-xylose to (D)-xylulose, the phosphorylation of (D)-xylulose to obtain (D)-xylulose-1-phosphate (Xyl1P), and the aldolase cleavage of the latter to yield glycolaldehyde and DHAP. Both compounds can be further processed via the annotated natural metabolic network. Stoichiometric analyses showed that the synthetic pathway provides an efficient access to the value-added two-carbon (C2) compounds ethylene glycol and glycolic acid, which are either inaccessible via the annotated metabolic network of E. coli or produced at 20 % lower theoretical yield, respectively. The simultaneous expression of xylulose-1 kinase and Xyl1P aldolase activities, which were provided by human ketohexokinase-C and human aldolase-B, respectively, was necessary and sufficient to restore growth of a (D)-xylulose-5-kinase (?xylB) mutant on xylose. In this strain, ethylene glycol was the major metabolic end-product and produced at a molar yield of 0.47. Further metabolic engineering provided strains that assimilated the entire C2 fraction back into the central metabolism, or that produced 4.3 g/l glycolic acid at a molar yield of 0.9 in shake flasks. The wild-type (WT) culture and synthetic strain (PEN205) were used for the microarray analysis. PEN205 is a ?xylB deleted strain that express the synthetic pathway. WT and PEN205 were grown to the logarithmic phase of growth in M9 minimal medium supplemented with xylose. PEN205 culture RNA was extracted when their OD was around 1. WT culture were split into two equal aliquots at the OD of ~1 and further cultivated in the presence or absence of 10 mM glycolaldehyde. After 30 min of incubation, cells were used for RNA extraction as well.
2015-05-27 | E-GEOD-69265 | biostudies-arrayexpress
Project description:Non-alcoholic fatty liver disease (NAFLD) is a major public health burden and it covers a spectrum of diseases. NAFLD starts with the accumulation of lipid droplets (LDs) within hepatocytes (steatosis). Part of the challenge of studying the mechanistic processes involved in LD accumulation and their implications on the pathogenesis of human NAFLD is due to the available models. Investigating hepatic LDs in humans is challenging and relies on liver biopsies, meaning only cross-sectional data be obtained. On the other hand, LD patterns in in vitro models are poorly defined and rarely reported. Diacylgylcerol acyltransferase (DGAT)2 is one of two enzymes that carry out the final committed step in triacylglycerol (TAG) synthesis. It is unclear whether the enzymes are able to compensate for each other or whether they have distinct roles. It has been hypothesised that DGAT1 primarily utilises exogenous fatty acids and DGAT2 uses de novo-derived fatty acids. Given the important role of this enzyme in TAG synthesis and accumulation, the aims of this study are first to create a cellular model of intrahepatocellular TAG accumulation by manipulating nutritional substrates and to investigate intracellular metabolism in wildtype and DGAT2 knockout cells under these conditions. The experimental workflow for this study is as follows: Huh7 cells (either wild type or knockout) were grown in media containing 11 mM glucose and 2% human serum (HS) for seven days before additional sugars and fatty acids (FAs) were added for a further seven days. All treatments contained 11 mM glucose and 2% HS, either with 200 µM FAs (low fat low sugar; LFLS), 5.5 mM fructose + 200 µM FAs (low fat high sugar; LFHS) or 5.5 mM fructose + 800 µM FAs (high fat high sugar; HFHS). FA metabolism, lipid droplet characteristics and transcriptomic signatures were investigated.
2020-08-21 | E-MTAB-7884 | biostudies-arrayexpress