Comparison of gene expression of WT and PerM mutant M. tuberculosis H37Rv grown in high and reduced magnesium
ABSTRACT: M. tuberculosis membrane protein PerM is required for persistence in chronic mouse infection. In vitro, the perM mutant requires increased magnesium for replication and survival, with elongation and lysis in low-magnesium media. To gain insight into the function of PerM, we compared the transcriptomes of wt and perM mutant Mtb grown in high (2000 μM) and reduced (250 μM) Mg2+ in three independent experiments. Transcriptome analysis of the perM mutant grown in reduced magnesium revealed upregulation of cell division and cell wall biosynthesis genes. Bacteria were grown for 5 days in Sauton’s media containing 250 or 2000 μM MgCl2 and 0.05% Tween 80. Three independent experiments were performed.
Project description:In our previous work, we showed the positive effect of the magnesium and the negative effect of the copper on yeast fermentation performance. The magnesium increases the ethanol yield and a faster glucose consumption by the yeast, on the other hand, the copper provides an opposite effect in yeast under fermentation condition. Therefore, from this contrasting effect we performed the gene-wide expression analysis in the industrial yeast Saccharomyces cerevisiae JP1 under fermentation condition in order to reveal the gene expression profile upon magnesium and copper supplementation. Fermentation assays was performed with the industrial yeast S. cerevisiae JP1 in reference medium (mineral concentration balanced), in the medium supplemented with 500 mg/L of magnesium (Mg2+ medium) and in the medium supplemented with 1 mg/L of copper (Cu2+ medium).
Project description:To understand the Abscisic Acid (ABA) signaling in response to dehydration stress, we performed analysis of gene expression using Arabidopsis wild-type plants and the nced3-2 mutant under dehydration stress. The nced3-2 mutant is an Arabidopsis T-DNA tagged knock-out mutant of the NCED3 gene, which has an essential role in dehydration-inducible ABA biosynthesis. Arabidopsis plants were grown in in soil (verdenite 40 mmΦ, Verde Co., Ltd., Kanagawa, Japan) in a cell strainer (Falcon, 40 μm; Corning Inc., NY, USA). Plants were grown at 22°C for 3 weeks under illumination (40–60 μmol m-2s-1; 16 h light/8 h darkness). Three-week-old plants were exposed to dehydration stress by being denied water for 6, 24, 48, or 72 h.
Project description:M. tuberculosis thioredoxin reductase (TrxB2) is essential for bacterial survival in vitro and in vivo. To gain insight into the mechanisms underlying lethality caused by TrxB2 depletion, we compared the mRNA profiles of TrxB2-DUC mutant in the presence and absence of atc. We grew bacteria in 7H9 medium to an OD of 0.5~0.6 and then added atc to a final concentration of 400ng/ml. Samples were collected 6 h and 24 h after atc treatment.
Project description:M. tuberculosis thioredoxin reductase (TrxB2) is essential for bacterial survival in vitro and in vivo. To gain insight into the pathways dependent on TrxB2, we compared the mRNA profiles of TrxB2-DUC mutant in the presence and absence of atc and DTT. We grew bacteria in 7H9 medium to an OD of 0.5~0.6. We then added atc to a final concentration of 800 ng/ml and DTT to a final concentration of 2 mM. Samples were collected 24 hr after treatment.
Project description:Martian regolith (unconsolidated surface material) is a potential medium for plant growth in bioregenerative life support systems during manned missions on Mars. However, hydrated magnesium sulfate mineral levels in the regolith of Mars can reach as high as 10 wt%, and would be expected to be highly inhibitory to plant growth. A global approach was used to identify novel genes with potential to enhance tolerance to high MgSO4 stress. The early Arabidopsis root transcriptome response to elevated concentrations of magnesium sulfate was characterized in col-0, and also between col-0 and the mutant line cax1-1 – a mutant relatively tolerant of high levels of MgSO4•7H2O in soil solution. After 3 weeks of growth under hydroponic conditions, Arabidopsis thaliana col-0 roots were exposed to a basic nutrient solution (0.25 g/L MES, 1/16x MS, pH 5.7) with an additional 2.08 mM magnesium sulfate (total Ca:Mg ratio = 1:15) for 45 min., 90 min., or 180 min., while a col-0 control set was exposed to the basic nutrient solution without additional magnesium sulfate for 45 minutes. Arabidopsis thaliana cax1-1 roots were exposed to the basic nutrient solution with additional magnesium sulfate for 180 min. only. Four replicate containers were harvested for the control and each of the treatment sets, resulting in a total of 20 samples. Gene expression of the col-0 sets exposed to magnesium sulfate treatment for 45 min., 90 min., or 180 min. was compared to gene expression of the col-0 control set. Gene expression of the cax1-1 set exposed to magnesium sulfate treatment for 180 min. was compared to gene expression of the col-0 set exposed to magnesium sulfate treatment for 180 minutes.
Project description:To gain insight into the functional mechanisms of an RXR agonist HX630 in murine pituitary corticotroph tumor cells (AtT20 cell), especially concerning cell proliferation and apoptosis, we conducted whole genome microarray analysis to identify gene expression patterns, networks, and pathways in AtT20 cells treated with HX630 at 10 μM for 48 hr. 2,173 up-regulated and 3,169 down-regulated genes were identified that demonstrated fold-changes of at least 1.5 and a P-value <0.01 in AtT20 cells with HX630 at 10 uM compared with control. Both the up- and down-regulated differentially expressed genes were submitted to IPA, which groups significant genes according to biological processes in which they function, and identifies various functions, networks, and pathways. These analyses showed that the categories of the biological functions and diseases significantly regulated by HX630 were associated with cell death, growth and cancer. Furthermore, it was identified that a gene network associated with caspase 3 was significantly induced by HX630.These data indicated that HX630 induced cell death and suppressed cell growth in AtT20 cells, consistent with the results of in vitro studies. AtT20 cells grown to 50 % confluence in regular medium in 12-multiwell plates were incubated either without or with HX630 at 10 μM in DMEM supplemented with 1 % stripped FBS media for 48 hr. The cells were then lysed and their total RNAs were isolated for microarray analysis.
Project description:Physiological mechanisms involved in root hair development in response to magnesium (Mg) availability are unclear. This study investigated the influence of Mg availability on root hair development in arabidopsis grown in different Mg concentrations ranging from 0.5 μM to 10 mM. After 7-d treatment, root hair development was enhanced in roots exposed to low Mg but was inhibited severely in roots grown in high Mg. Low Mg (0.5 µM) enhanced many genes like LRX1, COW1, EXP7 and ROP2 that control root hair development. Low Mg supply also increased concentrations of total Ca2+ and ROS in roots, but application of either BAPTA or DPI to low Mg treatment blocked the enhanced development of root hairs. The opposite was true when the plants under high Mg (3 mM) were supplied with Ca2+ or PMS. Besides, in roots under low Mg and high Mg, the most significant biological function enrichment were in the ‘oxidation reduction’, ‘cell wall organization’, ‘ion response’. This study demonstrated that Ca2+ and ROS played critically in controlling the Mg-induced development of root hairs. Meanwhile, transcriptome analysis associated with Mg supply contributed to a better understanding of molecular events responsible for sensing Mg status. Roots were sampled from five-week-cultivated Arabidopsis after 7 d treatment of low Mg (supplied with 0.5 μM Mg2+) and high Mg (supplied with 10 mM Mg2+).
Project description:Transcriptional profiling of three sequenced S. enterica strains: S. Typhimurium LT2, S. Typhi CT18, and S. Typhi Ty2 in PhoP-inducing and non-inducing conditions in vitro, and compared these results to profiles of phoP-/Q- mutants derived from S. Typhimurium LT2 and S. Typhi Ty2. Two-condition experiment: Each strain above was grown in PhoP-inducing (Low Magnesium concentration) and PhoP non-inducing conditions (High Magnesium concentration) with 1 dye reversal.
Project description:This project focused in the proteomic characterization of the secreted proteome of a Rv1002c (Protein mannosyltransferase) knock-out mutant and its wild type counterpart. The analysis was carried out using a M. tuberculosis ΔleuDΔpanCD auxotroph Rv1002c wild type compared to M. tuberculosis ΔleuDΔpanCDΔRv1002c. Both strains were grown in triplicate and their whole cell lysate (WCL) and secreted proteins (CFP) where purified, trypsin digested and characterized by liquid-chromatography coupled with tandem mass spectrometry (LC-MS/MS). Three replicate injections were performed for each of the sampels. The resulting spectra were searched against an M. tuberculosis database and relative protein abundance was determined using normalized spectral counts. Protein abundance differences were tested by Student’s t test. Abundance of 39 secreted proteins was significantly different between the WT and the ΔRv1002c. Decrease abundance of known glycoproteins, including several lipoglycoproteins was observed in the ΔRv1002c strain. In the WCL, 61 proteins presented significantly different abundance between the WT and the ΔRv1002c strains, including 4 proteins that were completely absent in the mutant (Rv2510c, PknF, Mpt70 and CeoC).
Project description:The purpose of this study was to examine how ethoxzolamide modulates gene M. tuberculosis gene expression at acidic pH. We observed that ethoxzolamide downregulates genes of the PhoPR regulon. Mtb strain CDC1551 was grown at 37C in T-25 vented, standing tissue culture flasks in 8 ml of buffered 7H9 medium seeded an initial OD of 0.2. Three conditions were examined with two biological replicates: 1) DMSO treated, pH 7.0, 2) DMSO treated, pH 5.7, 3) 40 µM ethoxzolamide treated, pH 5.7. The CDC1551(phoP::Tn) mutant was grown in a similar manner and treated with DMSO at pH 7.0 and 5.7. Following six days incubation, total bacterial RNA was extracted and analyzed as previously described.