Effect of Calcium Chloride Concentrations on the Transcriptome of Brevibacterium lines
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ABSTRACT:
ORGANISM(S): Brevibacterium linens
PROVIDER: GSE93668 | GEO | 2020/01/16
REPOSITORIES: GEO
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Project description:Purpose: Compare the transcriptional profile of staurosporine-treated cells in Neurospora crassa wild type cells grown with different availabilities of calcium. Methods: Conidial suspensions were obtained and 1 x 106 cells/ml incubated in minimal medium (with the indicated concentrations of calcium) for 6 hours (26ºC, 140 rpm, constant light) followed by the addition of staurosporine (or DMSO) and growth for 1 more hour. For the experiments in 0 Ca2+ or 20 mM CaCl2 media, the concentration of KH2PO4 in the 50x Vogel’s stock solution was limited to 10 mM to avoid precipitation with supplemental calcium. Cells were harvested using 0.45 μm filters and immediately frozen in liquid nitrogen. Total RNA was isolated by the Trizol-Phenol-Chloroform method. After digestion of 25 μg RNA with TURBO DNAse (Life Technologies), mRNA was purified using Dynabeads oligo(dT) magnetic beads (Life Technologies). The mRNA was chemically fragmented using the Ambion RNA fragmentation kit (Life Technologies). First and second strand cDNA synthesis was achieved using appropriate kits (Life Technologies). The illumina TruSeq kit was employed to generate the cDNA libraries with indexing adapters essentially following the manufacturer’s protocol. After purification of the libraries with AMPure XP beads (Roche), the quality of the libraries was analyzed in a Agilent 2100 Bioanalyzer. The cDNA libraries were sequenced in a illumina HiSeq2000 and single reads of 50 bp were obtained. Sequencing data was handled essentially with Tophat, Cufflinks and Cuffdiff. Expression levels are presented as Fragments Per Kilobase of transcript per Million mapped reads (FPKM). Results: Differences in the concentration of extracellularly available Ca2+ result in distinct transcriptional programs. Conclusions: Our transcriptomic analysis provides a reference dataset for future investigations on the role of Ca2+ in fungal biology.
2014-12-05 | GSE53013 | GEO
Project description:Purpose: Compare the transcriptional profile of staurosporine-treated cells in Neurospora crassa wild type cells grown with different availabilities of calcium. Methods: Conidial suspensions were obtained and 1 x 106 cells/ml incubated in minimal medium (with the indicated concentrations of calcium) for 6 hours (26M-BM-:C, 140 rpm, constant light) followed by the addition of staurosporine (or DMSO) and growth for 1 more hour. For the experiments in 0 Ca2+ or 20 mM CaCl2 media, the concentration of KH2PO4 in the 50x VogelM-bM-^@M-^Ys stock solution was limited to 10 mM to avoid precipitation with supplemental calcium. Cells were harvested using 0.45 M-NM-<m filters and immediately frozen in liquid nitrogen. Total RNA was isolated by the Trizol-Phenol-Chloroform method. After digestion of 25 M-NM-<g RNA with TURBO DNAse (Life Technologies), mRNA was purified using Dynabeads oligo(dT) magnetic beads (Life Technologies). The mRNA was chemically fragmented using the Ambion RNA fragmentation kit (Life Technologies). First and second strand cDNA synthesis was achieved using appropriate kits (Life Technologies). The illumina TruSeq kit was employed to generate the cDNA libraries with indexing adapters essentially following the manufacturerM-bM-^@M-^Ys protocol. After purification of the libraries with AMPure XP beads (Roche), the quality of the libraries was analyzed in a Agilent 2100 Bioanalyzer. The cDNA libraries were sequenced in a illumina HiSeq2000 and single reads of 50 bp were obtained. Sequencing data was handled essentially with Tophat, Cufflinks and Cuffdiff. Expression levels are presented as Fragments Per Kilobase of transcript per Million mapped reads (FPKM). Results: Differences in the concentration of extracellularly available Ca2+ result in distinct transcriptional programs. Conclusions: Our transcriptomic analysis provides a reference dataset for future investigations on the role of Ca2+ in fungal biology. Transcriptional profile of staurosporine-treated Neurospora crassa wild type cells growing with different concentrations of calcium was compared using illumina HiSeq2000 and single reads of 50 bp.
2014-12-05 | E-GEOD-53013 | biostudies-arrayexpress
Project description:Cell surface calcium (Ca2+) channels permit Ca2+ ion influx, with Ca2+ taking part in cellular functions such as proliferation, survival, and activation. The expression of voltage-dependent Ca2+ (CaV) channels may modulate the growth of hematologic cancers. Profile analysis of Ca2+ channels, with a focus on the L-type CaV channels, was performed on RNA sequencing data from lymphoma and leukemia cell lines and samples derived from patients with diffuse large B cell lymphoma (DLBCL). CaV1.2 expression was found to be elevated in classical Hodgkin lymphoma (CHL) cell lines when compared to other B cell lymphoma cell lines. In our analysis comparing activated B cell-like DLBCL (ABC-DLBCL) and germinal centre B cell-like DLBCL (GCB-DLBCL) patient samples, ABC-DLBCL revealed stronger expression of CaV1.3, whereas CaV1.1, CaV1.2, and CaV1.4 showed greater expression levels in GCB-DLBCL. As Ca2+ is known to bind to calmodulin, leading to calcineurin activation and the passage of nuclear factor of activated T cells (NFAT) to the cell nucleus, pathways for calcineurin, calmodulin, NFAT, and Ca2+ signaling were also analyzed by gene set enrichment analysis. The NFAT and Ca2+ signaling pathways were found to be upregulated in the CHL cell lines relative to other B cell lymphoma cell lines. Furthermore, the calmodulin and Ca2+ signaling pathways were shown to be downregulated in the ABC-DLBCL patient samples. The findings of this study suggest that L-type CaV channels and Ca2+-related pathways could serve as differentiating components for biologic therapies to target hematological cancers.
2022-06-15 | GSE189927 | GEO
Project description:TRPM3 belongs to the melastatin sub-family of transient receptor potential (TRPM) cation channels and has been shown to function as a steroid-activated, heat-sensitive, calcium ion (Ca2+) channel. A missense substitution (p.I65M) in the TRPM3 gene of humans (TRPM3) and mice (Trpm3) has been shown to underlie an inherited form of early-onset, progressive cataract. Here we model the pathogenetic effects of this cataract-causing mutation using ‘knock-in’ mutant mice and human cell-lines. Trpm3 and its intron-hosted micro-RNA gene (Mir204) were strongly co-expressed in the lens epithelium and other non-pigmented and pigmented ocular epithelia. Homozygous Trpm3-mutant lenses displayed elevated cytosolic Ca2+ levels and an imbalance of sodium (Na+) and potassium (K+) ions coupled with increased water content. Homozygous TRPM3-mutant human lens epithelial (HLE-B3) cell-lines and Trpm3-mutant lenses exhibited increased levels of phosphorylated mitogen-activated protein kinase 1/extracellular signal-regulated kinase 2 (MAPK1/ERK2/p42) and MAPK3/ERK1/p44. Mutant TRPM3-M65 channels displayed an increased sensitivity to external Ca2+ concentration and an altered dose response to pregnenolone sulfate (PS) activation. Trpm3-mutant lenses shared downregulation of genes involved in insulin/peptide secretion and upregulation of genes involved in Ca2+ dynamics. By contrast, Trpm3-deficient lenses did not replicate the pathophysiological changes observed in Trpm3-mutant lenses. Collectively, our data suggest that a cataract-causing substitution in the TRPM3 cation channel elicits a deleterious gain-of-function, rather than a loss-of-function, mechanism in the lens.
2024-02-14 | GSE250518 | GEO
Project description:Catheter-associated urinary tract infections (CAUTIs) account for over 30% of acute nosocomial infections in the U.S. and generate $340 million in healthcare costs annually. A major causative agent of CAUTIs is Proteus mirabilis, an understudied Gram-negative pathogen noted for its ability to form urinary stones via the activity of urease. Urease mutants cannot induce stones and are attenuated in a murine UTI model, indicating this enzyme is essential to P. mirabilis pathogenesis. The ability to induce urinary stone formation requires an active urease, a nickel metalloenzyme that hydrolyzes urea. This reaction produces ammonia as a byproduct, which can serve as a nitrogen source and weak base that raises the local pH. The resulting alkalinity induces the precipitation of polyvalent cations and anions to form stones. Expression of urease genes is activated by transcriptional regulator UreR in a urea-dependent manner. Thus, urease genes are highly expressed in the urinary tract where urea is abundant (~400 mM in human urine). Production of mature urease also requires the import of nickel into the cytoplasm and its incorporation into the urease apoenzyme. Urease accessory proteins primarily acquire nickel from the Ynt transporter and facilitate the incorporation of nickel to form mature urease. P. mirabilis encodes a second, low-affinity transport system (Nik) that can provide nickel when this metal is abundant. In this study, we identified UreR as the first defined regulator of nickel transport in P. mirabilis. We also offer evidence for direct regulation of the ynt promoter by UreR. Using bioinformatics, we identified UreR-regulated urease loci in 15 Morganellaceae family species across three genera. Additionally, we located two mobilized UreR-regulated urease loci that also encode the ynt transporter, implying that UreR regulation of ynt is a conserved regulatory relationship. Our study demonstrates that UreR regulates all genes required to produce mature urease, an essential virulence factor for P. mirabilis uropathogenesis.
2024-03-25 | GSE254779 | GEO
Project description:The response of osteoprogenitors to calcium (Ca2+) is of primary interest for both normal bone homeostasis and the clinical field of bone regeneration. The latter makes use of calcium phosphate-based bone void fillers to heal bone defects, but it is currently not known how Ca2+ released from these ceramic materials influences cells in situ. Here, we have created an in vitro environment with high extracellular Ca2+ concentration and investigated the response of human bone marrow-derived mesenchymal stromal cells (hMSCs) to it. Ca2+ enhanced proliferation and morphological changes in hMSCs. Moreover, the expression of osteogenic genes is highly increased. A 3-fold up-regulation of BMP-2 is observed after only 6 h and pharmaceutical interference with a number of proteins involved in Ca2+ sensing showed that not the calcium sensing receptor, but rather type L voltage-gated calcium channels are involved in mediating the signaling pathway between extracellular Ca2+ and BMP-2 expression. MEK1/2 activity is essential for the effect of Ca2+ and using microarray analysis, we have identified c-Fos as an early Ca2+ response gene. We have demonstrated that hMSC osteogenesis can be induced via extracellular Ca2+, a simple and economic way of priming hMSCs for bone tissue engineering applications. For more information check: https://cbit.maastrichtuniversity.nl/
2018-10-06 | E-MTAB-7222 | biostudies-arrayexpress
Project description:Calcium acts as a universal second messenger to regulate gene expression in both developmental processes and responses to environmental stresses. Previous studies showed that a number of stimuli can induce calcium increases in the cytoplasm and nucleus, independently. However, the gene expression network deciphering [Ca2+]cyt and/or [Ca2+]nuc signaling pathway remain obscure. Using transgenic Arabidopsis containing a fusion protein, comprising rat parvalbumin (PV) with either a nuclear export sequence (PV-NES) or a nuclear localization sequence (NLS-PV), to selectively buffer the cytosolic or nucleosolic calcium, we identified the [Ca2+]cyt- or /and [Ca2+]nuc-regulated ABA- and MeJA-responsible genes with the Arabidopsis Genome Oligo array.
2018-01-25 | GSE109611 | GEO
Project description:Calcium is an essential macronutrient and plant requires it in large amounts for normal growth and development. This ion participated in innumerable processes and affects nearly all aspects of plant growth and development such as signal transduction, metabolism of lipids, proteins, and carbohydrates, cell growth, cell wall and membrane stabilization. We used whole genome microarrays to determine the transcriptomic profile of rice seedlings exposed to short-term and long term Ca2+ deficiency followed by Ca2+ resupply. Calcium treated seedlings were used for for RNA extraction and hybridization on Agilent microarray platform. Three biological replicates of each sample were used for microarray analysis. We wanted to know the altered expression patterns of calcium-responsive genes majorly involved in metabolic processes, signal transdction pathways, transcriptional regulation, and transport of multiple molecules including Ca2+. Seeds of indica rice were surface sterilized and grown hydroponically in rice growing medium. Plants were grown in the rice growth room under the condition of 16 h light/8 h dark (28M-BM-0C) photoperiod with 70 % humidity. After 5 days of normal growth, some of the seedlings were transferred to nutrient solution lacking CaCl2 (-Ca2+). After 5 and 14 days, calcium (0.798 mM CaCl2) was resupplied to the plants grown in -Ca2+ medium for 6 h.
2014-01-17 | E-GEOD-50917 | biostudies-arrayexpress
Project description:In Saccharomyces cerevisiae, the Ca2+/calmodulin-dependent protein phosphatase, calcineurin, is activated by specific environmental conditions, including exposure to Ca2+ and Na+, and induces gene expression by regulating the Crz1p/Tcn1p transcription factor. We used DNA microarrays to perform a comprehensive analysis of calcineurin/Crz1p-dependent gene expression following addition of Ca2+ (200 mM) or Na+ (0.8 M) to yeast. 163 genes exhibited increased expression that was reduced 50% or more by calcineurin inhibition. These calcineurin dependent genes function in signaling pathways, ion/small molecule transport, cell wall maintenance, vesicular transport, and include many open reading frames of heretofore-unknown function. Three distinct gene classes were defined: 28 genes displayed calcineurin-dependent induction in response to Ca2+ and Na+, 125 showed calcineurin-dependent expression following Ca2+ but not Na+ addition, and 10 were regulated by calcineurin in response to Na+ but not Ca2+. Analysis of crz1D cells established Crz1p as the major effecter of calcineurin-regulated gene expression in yeast. We identified the Crz1p binding site as 5-GNGGC(G/T)CA-3 by in vitro site selection. A similar sequence, 5-GAGGCTG-3, was identified as a common sequence motif in the upstream regions of calcineurin/Crz1p-dependent genes. This finding is consistent with direct regulation of these genes by Crz1p.
2005-09-23 | E-SMDB-1427 | biostudies-arrayexpress
Project description:In Saccharomyces cerevisiae, the Ca2+/calmodulin-dependent protein phosphatase, calcineurin, is activated by specific environmental conditions, including exposure to Ca2+ and Na+, and induces gene expression by regulating the Crz1p/Tcn1p transcription factor. We used DNA microarrays to perform a comprehensive analysis of calcineurin/Crz1p-dependent gene expression following addition of Ca2+ (200 mM) or Na+ (0.8 M) to yeast. 163 genes exhibited increased expression that was reduced 50% or more by calcineurin inhibition. These calcineurin dependent genes function in signaling pathways, ion/small molecule transport, cell wall maintenance, vesicular transport, and include many open reading frames of heretofore-unknown function. Three distinct gene classes were defined: 28 genes displayed calcineurin-dependent induction in response to Ca2+ and Na+, 125 showed calcineurin-dependent expression following Ca2+ but not Na+ addition, and 10 were regulated by calcineurin in response to Na+ but not Ca2+. Analysis of crz1D cells established Crz1p as the major effecter of calcineurin-regulated gene expression in yeast. We identified the Crz1p binding site as 5-GNGGC(G/T)CA-3 by in vitro site selection. A similar sequence, 5-GAGGCTG-3, was identified as a common sequence motif in the upstream regions of calcineurin/Crz1p-dependent genes. This finding is consistent with direct regulation of these genes by Crz1p.
2013-01-30 | E-SMDB-15 | biostudies-arrayexpress