RNA-seq of SH-SY5Y NB cells stably transduced with shETV5 compared to parental cells, cell line samples and samples of xenografted tumors
Ontology highlight
ABSTRACT:
INSTRUMENT(S): NextSeq 500
ORGANISM(S): Homo sapiens
SUBMITTER: Liselot Mus
PROVIDER: E-MTAB-6713 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
Similar Datasets
Project description:SH-SY5Y cells were transfected with an siRNA against T-UC.300A (final concentration 50nM) or negative control siRNA (Dharmacon Negative Control #1, final concentration 50 nM) using the transfection reagent Lipofectamine (Invitrogen). Media was changed after 24hrs. RNA was extracted 120hrs after transfection. Gene expression microarray analysis was carried out using Roche NimbleGens 4x72K Homo Sapiens gene expression array.
2012-10-14 | E-MEXP-3712 | biostudies-arrayexpress
Project description:TCF4 is an important neurodevelopmental transcription factor associated with schizophrenia and Pitt-Hopkins syndrome. In this study, we used genome-wide expression profiling to determine the effects of acute TCF4 knockdown on gene expression in SH-SY5Y neuroblastoma cells. Pathway and enrichment analysis on the differentially expressed genes in TCF4-knockdown cells identified an over-representation of genes involved in TGF-β signaling, epithelial-to-mesenchymal transition (EMT) and apoptosis. Among the most significantly differentially expressed genes were the EMT regulators SNAI2 and DEC1 and the proneural genes NEUROG2 and ASCL1. Altered expression of several mental retardation genes such as UBE3A (AS), ZEB2 (MWS) and MEF2C was also found in TCF4-depleted cells. These data suggest that TCF4 regulates a number of convergent signaling pathways involved in cell differentiation and survival in addition to a subset of clinically important mental retardation genes. To identify TCF4-mediated gene expression changes in SH-SY5Y cells, experiments were conducted with two control (mock and GAPDH KD) and two TCF4 knockdown (KD1 and KD2) groups. Two non-overlapping siRNAs against TCF4 were designed and transfected over a 72h period to induce global knockdown of TCF4 transcripts. In parallel, cells were mock transfected (mock; transfection reagent only) and transfected with an unrelated siRNA against GAPDH (GAPDH KD) in order to control for background gene expression changes due to transfection, presence of dsRNA and activation of the cellular silencing machinery. After transfection, RNA was extracted from all cells, converted to cDNA and labelled for microarray hybridization. Each of the four treatment groups (mock, GAPDH KD, KD1 and KD2) contained three biological replicates which were processed at the same time using the Toray microarray platform.
2013-08-20 | E-GEOD-48367 | biostudies-arrayexpress
Project description:C57BL/6 mice were maintained in specific pathogen-free conditions in accordance with the Institutional Animal Care and Use Committee. Listeria monocytogenes (clinical isolate 10403s) was from V. Boyartchuk (NTNU, Trondheim, Norway) and infected as previously described (PMID 24477914) for 24 hours before harvesting the spleen. PBS was used as control. . Single cell suspensions from spleens were used to make total RNA. 4 μg of total RNA was used to generate libraries for RNA-sequencing (Illumina unstranded mRNA kits). Samples were sized, quantified and validated on a Bioanalyzer. Libraries were sequenced on a High-Seq System (Illumina 2000, San Diego, CA) as paired-end 100 reads. Sequence reads were aligned to the mouse genome (assembly NCBI m37/mm9) using TopHat.Gene level read counts based on the resulting alignments were calculated using HTSeq. The DESeq R package was used to normalize gene counts, calculate fold change in gene expression, estimate p-values and adjusted p-values for change in gene expression values, and to perform a variance stabilized transformation on read counts.
2015-06-30 | E-MTAB-3315 | biostudies-arrayexpress
Project description:Today, the pathogenesis of human enterovirus type 71 (HEV71) infection in human central neural system remains unclear. HEV71 is the major pathogen of hand-foot-and-mouth disease (HFMD), and has been associated with severe neurological disease and even death in infants and young children. We employed the human whole genome microarray analyze the mRNA profiling in human neuroblastoma cells SH-SY5Y infected with HEV71 after transfection. Firstly, SH-SY5Y cells were transfected wtih miR-1246 inhibitor and negtive control respectively using HiPerFect Transfection Reagent according to the manufacturer’s instructions. Then the cells were infected with HEV71 after transfection. After 12 hours infection, the cells were harvested to microarray analysis. The results showed the altered expression of mRNAs including up-regulated genes and down-regulated genes. Overall, this finding will help to understand the functional genes in HEV71-infected human neuroblastoma cells and miR-1246-virus-host interaction. SH-SY5Y cells were transfected wtih miR-1246 inhibitor and negtive control respectively using HiPerFect Transfection Reagent according to the manufacturer’s instructions. Then the cells were infected with HEV71 after transfection. After infection, the cells were harvested to microarray analysis. Total RNA of cells infected with HEV71 was extracted using the TRIZOL Reagent according to the manufacturer's instructions. Gene-expression profiling was performed for each pooling RNA sample separately on the GeneChip_ Porcine Genome Array at CapitalBio Corporation (Beijing, China).
2014-07-31 | E-GEOD-45817 | biostudies-arrayexpress
Project description:We conducted a proof-of-concept experiment to explore the possibility of using gene expression-based high throughput screening (GE-HTS) to find inhibitors of a signaling cascade, using platelet derived growth factor receptor (PDGFR) signaling as the example. To define ERK/PDGFR activation signature, we performed gene expression profiling using Affymetrix U133A arrays 0.5-4 hours following PDGF stimulation, thereby identifying those genes whose expression is correlated with PDGFR activity. In order to identify the component of the gene expression signature that was attributable to ERK activation by PDGFR (as opposed to other pathways downstream of PDGFR), we also pretreated the cells will the ERK inhibitors U0126 and PD98059, and repeated the gene expression profiling studies. Experiment Overall Design: To define ERK/PDGFR activation signature, SH-SY5Y cells were grown to confluence and starved overnight in serum-free medium in order to silence any sustained effects from growth factor signaling. Prior to induction with 50ng/ml PDGF, cells were treated with pathway inhibitors 74 mM Apigenin or 50 mM U0126, or with DMSO (carrier) for 60 minutes. Total RNA was isolated 30 minutes after PDGF addition. Experiments were performed in duplicate. The RNA was processed and hybridized with Affymetrix U133A GeneChips.
2008-04-06 | E-GEOD-7403 | biostudies-arrayexpress
Project description:transcriptional profiling of DYX1C1 perturbation experiments
2012-10-01 | E-MTAB-1128 | biostudies-arrayexpress
Project description:Presence of BM-infiltrating neuroblastoma (NB) cells occurs in about 50% of patients affected by this peculiar pediatric cancer. Children aged more than 18 months at diagnosis with metastatic NB are high-risk patients with 25-30% overall survival at 5 years, despite multimodal aggressive therapy. Little is known about the metastatic process in NB patients. Hereby, we focused on miRNA expression profiles of BM-infiltrating as compared to those of primary NB tumors. For this purpose, we analyzed 22 BM-infiltrating NB cells, 22 primary NB tumors, and 4 paired BM-infiltrating and primary tumor specimens, all from patients with metastatic disease and aged > 18 months at diagnosis. Statistical analysis indicated that in human BM-infiltrating cells miR-659-3p was down-modulated as compared to primary tumors, in all sets of samples. Functional studies in HTLA and SH-SY5Y NB cell lines by miR-659-3p mimic and inhibitors demonstrated that miR-659-3p regulates the expression of the transcription factor CNOT1, that in turn regulates the expression of AU-rich element (ARE)-containing target genes AKT3, BCL2, THSB2 and CYR61, all belonging to the focal adhesion pathway. Indeed, CNOT1 expression was found significantly higher, whereas that of the ARE-containing target genes was significantly lower in BM-infiltrating cells than in primary tumors. Our findings about a role of the focal adhesion pathway, regulated by miR-659-3p through CNOT1, in NB metastatic process are intriguing for the development of new therapeutic strategies aimed to interrupt the metastatic process. To evaluate the effect of miR-659-3p up- and down-regulation on gene expression, HTLA-230 and SH-SY5Y cells were transfected with specific miR-659-3p mirVana™ mimic and inhibitor, respectively (Ambion, Life Technologies, Carlsbad, CA, USA, catalog# MC and MH 11582). Samples transfected with mirVana™ miRNA Mimic Negative Control #1 and mirVana™ miRNA Inhibitor Negative Control #1 were used as reference conditions. Transfection was performed at 20 nM miRNA concentration in OptiMEM© medium (Sigma-Aldrich), using Lipofectamin RNAiMAX© (Life Technologies), according to manufacturer’s protocol. Cells were then cultured for 48 hours, checked for viability and processed for total mRNA and miRNA extraction, as described below. Transfection experiments were performed twice with similar results. HTLA-230 (kindly donated by Dr E. Bogenman, Los Angeles, USA) and SH-SY5Y (purchased from Banca Biologica and Cell Factory, Genoa, Italy) NB cell lines were cultured in RPMI 1640 medium supplemented with 10% heat inactivated FCS, 50 mg/ml streptomycin, 50 mg/ml penicillin and 2 mM glutamine (Sigma-Aldrich, Milan, Italy). The HTLA-230 cells present MYCN amplification and SH-SY5Y cells have normal MYCN status. Both cell lines were checked for MYCN amplification, morphology, proliferation rate and mycoplasma contamination, after thawing and within four passages in culture. To evaluate the effect of miR-659-3p up- and down-regulation on gene expression, 1x106 HTLA-230 and SH-SY5Y cells were transfected with specific miR-659-3p mirVana™ mimic and inhibitor, respectively (Ambion, Life Technologies, Carlsbad, CA, USA, catalog# MC and MH 11582). Samples transfected with mirVana™ miRNA Mimic Negative Control #1 and mirVana™ miRNA Inhibitor Negative Control #1 were used as reference conditions. Transfection was performed at 20 nM miRNA concentration in OptiMEM© medium (Sigma-Aldrich), using Lipofectamin RNAiMAX© (Life Technologies), according to manufacturer’s protocol. Cells were then cultured for 48 hours, checked for viability and processed for total mRNA and miRNA extraction, as described below. Transfection experiments were performed twice with similar results.
2015-04-20 | E-GEOD-65153 | biostudies-arrayexpress
Project description:This experiment captures expression over 60,000 well-annotated RefSeq human transcripts over RNA samples from SH-SY5Y neuroblastoma cells transfected with human and non-human primate microRNA mimic variants of miR-299-3p, miR-503-3p, miR-508-3p and miR-541-3p, as well as a RNA duplex negative control (C2 mimic, Dharmacon).
2016-04-03 | E-MTAB-3617 | biostudies-arrayexpress
Project description:S-palmitoylation is the covalent attachment of palmitic acid to cysteine residues through a thioester bond. S-palmitoylation is highly abundant in neurons, where it plays a fundamental role in neuronal development. In addition, S-palmitoylation is associated with many neurodegenerative diseases, including Alzheimer's disease, Parkinson's disease, and Huntington's disease. However, knowledge of S-palmitoylation in neurodevelopment is limited due to technological challenges in analysing this highly hydrophobic modification of proteins. Here, we use two orthogonal methods, acyl-biotin exchange and lipid metabolic labelling, to identify S-acylated proteins and sites in neuronal cells. We identified 650 S-palmitoylated proteins by both methods, including proteins that are well-known for their role in neuronal differentiation. Importantly, significant changes in the abundance of S-palmitoylated proteins were detected during neuronal differentiation. Overall, the combination of methods described here provides the advantage of cross-validation of the identified S-palmitoylated proteins and should be used further to study S-palmitoylation in the central nervous system to understand physiology and neurodegenerative diseases.
2023-10-24 | PXD040613 | Pride
Project description:Background: SH-SY5Y cells exhibit a neuronal phenotype when treated with all-trans retinoic acid (RA), but the molecular mechanism of activation in the signaling pathway mediated by phosphatidylinositol 3-kinase (PI3K) is not sufficiently understood. To shed new light on the mechanism, we comprehensively compared the gene expression profiles between SK-N-SH cells and two subtypes of SH-SY5Y cells (SH-SY5Y-A and SH-SY5Y-E), each of which showed a different phenotype during RA-mediated differentiation. Results: SH-SY5Y-A cells differentiated in the presence of RA, whereas RA-treated SH-SY5Y-E cells required additional treatment with brain-derived neurotrophic factor (BDNF) for full differentiation. In combination with perturbation using a PI3K inhibitor, LY294002, we identified 386 genes and categorized them into two clusters dependent on the PI3K signaling pathway during RA-mediated differentiation in SH-SY5Y-A cells. Transcriptional regulation of the gene cluster was greatly reduced in SK-N-SH cells or partially impaired in SH-SY5Y-E cells in coincidence with a defect in the neuronal phenotype of these cell lines. Additional stimulation with BDNF induced a set of neural genes which were down-regulated in RA-treated SH-SY5Y-E cells but were abundant in the differentiated SH-SY5Y-A cells. Conclusions: We identified the gene clusters controlled by PI3K- and TRKB-mediated signaling pathways during differentiation in two subtypes of SH-SY5Y cells. TRKB-mediated bypass pathway compensates for the impaired neural functions generated by defects in several signaling pathways including PI3K in SH-SY5Y-E cells. The expression profiling data are useful for further studies to elucidate the signal transduction-transcriptional network including PI3K and/or TRKB. Experiment Overall Design: Human neuroblastomas, SK-N-SH (HTB-11) and SH-SY5Y-A cells (CRL-2266) were obtained from the American Type Culture Collection (ATCC). We also obtained SH-SY5Y-E cells (EC94030304) from the European Collection of Cell Cultures (ECACC). Tissue culture cells were maintained in D-MEM/F12 1:1 mixture supplemented with 15% FBS (Fetal Bovine Serum) and 1% NEAA (Non-essential amino acid) in a 5% CO2 humidified incubator at 37oC. The culture medium was changed twice a week. For the RA-inducible experiment, random culture cells from two clone subtypes of SH-SY5Y and SK-N-SH were seeded in laminin coated culture dishes (BioCoat Laminin Cellware; BD Biosciences, Billerica, MA, USA) for 1 day and then transferred to a medium containing 10 μM of RA in the presence or the absence of LY294002 (10μM) for five days. For BDNF-induced sequential differentiation of the SH-SY5Y-E strain, cells were washed with D-MEM/F12 twice after five days in the presence of RA and then incubated with 50 ng/ml of BDNF in D-MEM/F12 without serum for three days.
2009-09-04 | E-GEOD-9169 | biostudies-arrayexpress