The effect of ERG overexpression and ERG knockdown by shRNA in RWPE1 and VCAP cells, respectively
ABSTRACT: The goal of this project was to analyze the global gene expression profiles of RWPE1 and VCAP cells following transfection of GFP, GFP-ERG at 48 and 72hrs time points and stable ERG shRNA, scramble shRNA, respectively. Overall design: RWPE1 cells were transfected with GFP or GFP-ERG. VCAP cells were transfected with ERG lenti-shRNA or scramble shRNA. Transfections were performed in duplicate. Total cellular RNA was isolated with Trizol and quality analysed by the bioanalyser kit.
Project description:The goal of this project was to analyze the global gene expression profiles of RWPE1 and VCAP cells following transfection of GFP, GFP-ERG at 48 and 72hrs time points and stable ERG shRNA, scramble shRNA, respectively. RWPE1 cells were transfected with GFP or GFP-ERG. VCAP cells were transfected with ERG lenti-shRNA or scramble shRNA. Transfections were performed in duplicate. Total cellular RNA was isolated with Trizol and quality analysed by the bioanalyser kit.
Project description:Chromosomal rearrangements involving ETS factors, ERG and ETV1, occur frequently in prostate cancer. We here examine human prostate cancer cells control VCaP and LNCaP cells with ERG- or ETV1-silenced VCaP or LNCaP cells, respectively, in hormone deprived and stimulated conditions. VCAP and LNCaP cells, 24 hr after ERG or ETV1 RNA interference, respectively, were grown in hormone-depleted conditions for 2 days, and then in the presence of EtOH (vehicle) or 10nM DHT for 16hr. Total RNA was extracted from three biological replicates. This was used to hybridize to Affymetrix expression arrays using the HG-U133 Plus 2.0 platform.
Project description:VCaP cells expressing inducible shRNAs for either ERG or a non-targeting control were treated with Doxycycline for 1, 3, 7 and 10 days prior to collection This experiment is designed to see which genes and pathways are modulated by ERG knockdown VCaP cells stably expressing a Doxycycline (dox)-inducible control nontargeting shRNA (Pak4) or an ERG shRNA (2217) were exposed to 100ng/ml Dox for the noted days.
Project description:Microarray studies was performed to analyze gene expression profiling in VCaP cells after ERG was silenced by two different siRNAs or VCaP cells were treated by WP1130. RNA samples, containing 3 replicates for each sample, were extracted from VCaP cells at 72 hours siERG post-transfection or 24 hours WP1130 treatment. After biotin labeling, samples were hybrided on the Illumina Human HT12v4.0 Expression Beadchip
Project description:Replicates of ChIP seq data using AR and ERG antibodies from VCaP cell lines harboring inducible shRNA constructs for PRMT5 (3 independent, sh1, sh2, sh3) and 1 non targeting control (NTC), all sample stimulated with ligand, either DHT or R1881 Overall design: AR ChIPseq in VCaP cells, 3 replicates each of DHT stimulated cells with one of 4 inducible shRNA constructs, sh1, sh2, sh3 targeting PRMT5 or NTC non targeting controls. Furthermore 2 AR ChIPseq in the same system stimulated with R1881 for targeting constructs sh1, sh2 and NTC. ERG ChIPseq 2 replicates DHT stimulated, each of sh1, sh2, sh3 constructs. Cells where induced (doxycyclin) for shRNA expression on day0, hormone starved on day3 and harvested on day5.
Project description:The TMPRSS2-ERG fusion positive prostate cancer cell line VCaP was grown for 72h in steroid depleted media (RPMI 10% CDT FBS) prior to vehicle (ethanol) or androgen stimulation (1nM R1881). To allow detailed analysis of androgen regulated gene expression in the VCaP prostate cancer cell line total RNA was harvested every every hour up to 24h using trizol (Sigma), quantified using a Nanodrop spectrophotometer (ND-1000). RNA samples were prepared for analysis on Illumina Human 12 v3 BeadArrays. Raw bead-summary data were generated using GenomeStudio version 1.9.0 (Illumina Inc.). For integrative analysis expression array data were normalised in R using the beadarray software and BASH (see Supplementary Methods for details) (Cairns et al., 2008; Dunning et al., 2007).
Project description:We compared the genome occupancy for FLAG-tagged versions of the ETS factors ERG and EHF in the normal prostate epithelial cell line RWPE1. Our in vitro binding studies support a model whereby oncogenic ETS factors like ERG bind cooperativly with AP1 factors at closly spaced ETS-AP1 sites, while certain non-oncogenic factors like EHF bind anti-cooperatively with AP1 at the same sites. ETS and AP1 binding motifs were enriched in both ChIP datasets, but the ERG-FLAG bound reginos contained a much higher percentage of ETS-AP1 sites spaced in close proximity, consistent with our in vitro binding data. Overall design: ChIP-seq with anti-FLAG antibody in RWPE1 cells expressing either FLAG-ERG or FLAG-EHF
Project description:ERK activates ERG-mediated prostate cancer specific gene expression program by phopshorylation induced dissociation of PRC2 complex Overall design: ChIP-Seq of transcription factor ERG and its co-repressor EZH2 RNA-seq of polyA selected RNA from RWPE1 cells expressing either empty vector, ERG-WT, ERG-S96A and ERG-S96E
Project description:ERG activates prostate cancer specific gene expression program by recuriting RNA binding protein EWS Overall design: RNA-seq of polyA selected RNA from RWPE1 cells expressing either empty vector, ERG-WT, ERG-WT with EWS shRNA, and ERG-P436A