Transcription profiling of human GRE specific DNA-binding polyamide effect on dexamethasone-stimulated lung adenocarcinoma cell line
Ontology highlight
ABSTRACT:
ORGANISM(S): Homo sapiens
SUBMITTER: Peter Dervan
PROVIDER: E-GEOD-17307 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
Similar Datasets
Project description:Analysis of dexamethasone-stimulated A549 lung adenocarcinoma epithelial cells treated with a glucocorticoid response (GR) element (GRE) specific DNA binding polyamide. Polyamide designed to target the sequence 5'-WGWWCW-3' and disrupt GR-mediated gene expression. Effects of the GR antagonist mifepristone also examined.
2009-09-01 | GSE17307 | GEO
Project description:Androgen Receptor (AR) is essential for the growth and progression of prostate cancer in both hormone-sensitive and hormone-refractory disease. We have designed a sequence-specific DNA binding polyamide (1) that targets the consensus androgen response element (ARE). This polyamide binds the PSA promoter ARE, inhibits androgen-induced expression of PSA and several other AR-regulated genes in cultured prostate cancer cells, and reduces AR occupancy at the PSA promoter and enhancer. Down-regulation of PSA by this polyamide was comparable to that produced by the synthetic anti-androgen bicalutamide (Casodex) at the same concentration. Genome-wide expression analysis reveals that a similar number of transcripts are affected by treatment with the polyamide and with bicalutamide. Direct inhibition of AR-DNA binding by sequence-specific DNA binding small molecules could offer an alternative approach to antagonizing AR activity. A polyamide (2) that targets a different DNA sequence is included as a control. Experiment Overall Design: DHT (dihydrotestosterone)-stimulated LNCaP cells that were treatment with polyamide 1, polyamide 2, bicalutamide were compared to control cells that were also DHT-stimulated. Cells not stimulated with DHT were also compared to the DHT-stimulated controls. Three biological replicates were included for each treatment/condition except the no-DHT induced controls, which were in biological duplicate.
2009-04-23 | E-GEOD-7708 | biostudies-arrayexpress
Project description:Transcription mediated by hypoxia inducible factor (HIF-1) contributes to tumor angiogenesis and metastasis but is also involved in the activation of cell-death pathways and normal physiological processes. Given the complexity of HIF-1 signaling it could be advantageous to target a subset of HIF-1 effectors rather than the entire pathway. We compared the genome-wide effects of three molecules that each interfere with the HIF-1-DNA interaction: a polyamide targeted to the hypoxia response element (HRE), siRNA targeted to HIF-1α, and echinomycin, a DNA binding natural product with a similar but less specific sequence preference to the polyamide. The polyamide affects a subset of hypoxia-induced genes that are consistent with the binding site preferences of the polyamide. For comparison, siRNA targeted to HIF-1α and echinomycin each affect the expression of nearly every gene induced by hypoxia. Remarkably, the total number of genes affected by either polyamide or HIF-1α siRNA over a range of thresholds is comparable. The data shows how polyamides can be used to affect a subset of a pathway regulated by a transcription factor. In addition, this study offers a unique comparison of three complementary approaches towards exogenous control of endogenous gene expression. Experiment Overall Design: Hypoxia-mimetic DFO (deferoxamine)-stimulated U251 cells that were treated with polyamide 1, HIF-1α siRNA, and echinomycin were compared to control cells that were also DFO-stimulated. Cells not stimulated with DFO were also compared to the DFO-stimulated controls. Three biological replicates were included for each treatment/condition.
2008-06-15 | E-GEOD-7835 | biostudies-arrayexpress
Project description:Gene regulation by DNA binding small molecules could have important therapeutic applications. This study reports the investigation of a DNA-binding pyrrole-imidazole polyamide targeted to bind the DNA sequence 5M-bM-^@M-^Y-WGGWWW-3M-bM-^@M-^Y with reference to its potency in a subcutaneous xenograft tumor model. The molecule is capable of trafficking to the tumor site following subcutaneous injection and modulates transcription of select genes in vivo. A FITC-labeled analogue of this polyamide can be detected in tumor-derived cells by confocal microscopy. RNA deep sequencing (RNA-seq) of tumor tissue allowed the identification of further affected genes, a representative panel of which were interrogated by qRT-PCR and correlated with cell culture expression levels. Xenografts. Grafting with A549-luc-C8. Experiments were performed in female SCID-beige mice (Charles River) between 8 and 12 weeks of age. Cells were injected into the left flank area of the animals as suspensions of 25 x 106 mL-1 in RPMI, 200 M-BM-5L per injection. Treatment and tumor proliferation monitoring. Mice were treated following the schedule delineated in treatment protocol. Tumor proliferation was monitored using the XENOGEN imaging device. The animals were anesthetized with 2 5 % isoflurane and subsequently transferred to the imaging chamber, whereupon the isoflurane levels were reduced to 1-2.5 %. The floor of the imager was heated to +37 M-BM-:C to avoid hypothermia. Breathing frequency was monitored and not allowed to drop below 1 s-1, adjusting the isoflurane levels accordingly at all times. Endpoint criteria and euthanasia. Animal endpoint criteria encompassed weight loss of over 15 %, restriction of motor function by the engrafted tumor, dehydration of over 10 % and moribund behavior. Where appropriate, the animals were euthanized by asphyxiation in a CO2 chamber. Tumor tissue harvest. Animals were resected and tumors excised using standard forceps, scissors and surgical blades. The tumors were combined into one sample per condition and mechanically sheared in TRIZOL, employing a specialized device (tissue tearer, model 985370). Total RNA workup was performed following the standard TRIZOL procedure, followed by a DNAse digest.
2012-10-02 | E-GEOD-40218 | biostudies-arrayexpress
Project description:Nuclear factor kappaB (NF-kB) is a transcription factor that regulates various aspects of immune response, cell death and differentiation as well as cancer. In this study we introduce the Py-Im polyamide 1 that binds preferentially to the sequences 5'-WGGWWW-3' and 5' GGGWWW-3'. The compound is capable of binding to kB sites and reducing the expression of various NF-kB driven genes including IL6 and IL8 by qRT-PCR. Chromatin immunoprecipitation experiments demonstrate a reduction of p65 occupancy within the proximal promoters of those genes. Genome-wide expression analysis by RNA-seq compares the DNA-binding polyamide with the well-characterized NF-kB inhibitor PS1145, identifying overlaps and differences in affected gene groups and showing that both affect comparable numbers of TNFa inducible genes. Inhibition of NF-kB DNA binding via direct displacement of the transcription factor is a potential alternative to the existing antagonists. A549 cells were treated with either a Py-Im polyamide or PS1145, subsequently induced with TNFa and compared with the untreated TNFa induced and the basal state by RNAseq. The NF-kB binding motif was derived by ChIP-seq
2011-12-12 | E-GEOD-34329 | biostudies-arrayexpress
Project description:Prostate cancer is the most common cancer in men and AR downstream signalings promote prostate cancer cell proliferation. Oct1 is an AR interacting partner and regulates the transcriptional activity of AR. In order to investigate the Oct1 function in prostate cancer cells, we performed gene expression in AR-positive prostate cancer cell lines after siOct1 or pyrrole-imidazole (PI) polyamide targeting Oct1-binding treatment. We also treated cells with vehicle or androgen to analyze the effects of Oct1 on AR function. Observation of androgen dependent gene expression changes after treatment with siOct1 or polyamide targeting Oct1 with microarray.
2016-04-01 | E-GEOD-77650 | biostudies-arrayexpress
Project description:Gene expression array analysis component. Ligand-dependent transcription by the nuclear receptor glucocorticoid receptor (GR) is mediated by interactions with co-regulators. The role of these interactions in determining selective binding of GR to regulatory elements remains unclear. Recent findings indicate a large fraction of genomic GR binding coincides with chromatin that is accessible prior to hormone treatment, suggesting that receptor binding is dictated by proteins that maintain chromatin in an open state. Combining nucleolytic cleavage and chromatin immunoprecipitation with high-throughput sequencing, we identify the activator protein 1 (AP1) as a major partner for productive GR-chromatin interactions. AP1 is critical for GR-regulated transcription and recruitment to co-occupied regulatory elements, illustrating an extensive AP1-GR interaction network. Importantly, the maintenance of baseline chromatin accessibility facilitates GR recruitment and is dependent on AP1 binding. We propose a model where the basal occupancy of transcription factors act to prime chromatin and direct inducible transcription factors to select regions in the genome. Dexamethasone treatment of cells at zero hour untreated control and 4 hour treatment times, in the absence and presence of acidic-fos (A-fos), an AP1 dominant negative. A-fos expression is under tetracycline control (Tet-Off). This is part of a larger study: Transcription Factor AP1 Potentiates Chromatin Accessibility and Glucocorticoid Receptor Binding. There are 2 replicates per condition per time point.
2011-06-30 | E-GEOD-29983 | biostudies-arrayexpress
Project description:We report that a DNA minor groove binding hairpin pyrrole-imidazole (Py-Im) polyamide interferes with RNA polymerase II (RNAP2) activity in cell culture. Genome-wide mapping of RNAP2 binding shows reduction of occupancy preferentially at transcription start sites (TSS), while occupancy at enhancer sites are is unchanged. Genome-wide mapping of RNA polymerase II in LNCaP cells treated with DHT and DHT with Py-Im polyamide.
2013-01-31 | E-GEOD-43253 | biostudies-arrayexpress
Project description:To determine whether the polyamide-Chl conjugate 1R-Chl would cause similar changes in global gene expression in K562 cells, affymetrix gene chip analysis was performed using 1R-Chl. Through class comparison analysis, 1R-Chl affected the levels of transcription and genes of interest were determined. Experiment Overall Design: K562 cells were incubated with 1R-Chl (at 250 nM) or in the absence of polyamide, in triplicate for 24 h before RNA purification and microarray analysis at The Scripps Research Institute microarray facility. Affymetrix U133A Plus 2.0 GeneChips were hybridized in groups of three for each of the two groups. The Affymetrix probe set data were imported into BRB Arraytools (3.5.0 Beta 2), selecting the U133 chips used in the experiment and leaving all filters off.
2008-06-16 | E-GEOD-8832 | biostudies-arrayexpress
Project description:A1-2 cells treated with 100 nM Dexamethasone or ethanol vehicle for 8h. The ability of steroid hormone receptors to initiate a genetic program is tightly regulated by the chromatin environment of the responsive regions. Using the glucocorticoid receptor (GR) as a model factor for transcriptional initiation, we classified chromatin structure through Formaldehyde Assisted Isolation of Regulatory Elements (FAIRE), a technique designed to identify regulatory regions and open chromatin within the genome. We looked at dynamic changes in FAIRE signals prior to and following activation of GR with its ligand, dexamethesone, specifically at regions of receptor interaction. We have found a distribution of GR responsive regions that respond to activation by varying degrees of chromatin modulation. The majority of regions that demonstrate GR interaction also demonstrate increases in FAIRE signal in response to ligand. Most of these GR responsive regions fell within a narrow window of FAIRE signal in the basal chromatin state, suggesting a preferred chromatin structure for GR recruitment. Supporting this notion, global FAIRE-seq data indicated an enrichment of signal surrounding the GR binding site prior to activation. FAIRE signal induction correlated to an increase in nuclease sensitivity and overall FAIRE signal also represented a general accessibility of the chromatin. Further investigation into the requirement of ATPase-dependent chromatin remodeling showed response element specific effects of Brg-1 knockdown. FAIRE induction was universally decreased by Brg-1 depletion, but to varying degrees in a target specific manner. Taken together, these data suggest classes of nuclear receptor response regions that react to activation through different chromatin regulatory events and identify a novel identifier of chromatin structure that classifies the majority of response elements tested. Experiment performed in triplicate. 3 biological replicates of A1-2 cells treated with dexamethasone and 3 biological replicates of A1-2 cells treated with ethanol
2012-02-28 | E-GEOD-30592 | biostudies-arrayexpress