UC San Diego Friedmann Kang Neuronal Differentiation DSD3 SNM
Ontology highlight
ABSTRACT:
ORGANISM(S): Mus musculus
SUBMITTER: Tae Hyuk Kang
PROVIDER: E-GEOD-42662 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
Similar Datasets
Project description:The overall gene expression pattern of HPRT knockdown murine D3 stem cells during neuronal differentiation was invesitigated by microarray based gene expression analysis and the results were interpreted by GO, GSEA and signaling pathway analyses with GeneSpring GX and PANTHER Classification System. For microarray experiment, total RNAs from replicated HPRT knockdown and control cultures were pooled and the microarray experiment with each pooled sample was triplicated, respectively.
2012-11-22 | E-GEOD-42453 | biostudies-arrayexpress
Project description:To characterize mechanisms responsible for the CNS dopamine deficiency and the resulting neuropathology caused by deficiency of the housekeeping purine salvage function hypoxanthine guanine phospho- ribosyltransferase (HPRT) in the Lesch Nyhan Disease (LND), we have used microarray-based methods of global gene expression together with quantitative PCR and Western blot analysis to identify dysregulation of genes and aberrant cellular processes in human fibroblasts and in SH-SY5Y neuroblastoma cells made HPRT-deficient by transduction with a retrovirus stably expressing an shRNA targeted against HPRT. Analysis of the microarray expression data by Gene ontology (GO) and Gene Set Enrichment Analysis (GSEA) as well as by GeneSpring GX10 and Panther Classification System reveal that HPRT deficiency is accompanied by aberrations in a variety of pathways known to regulate neurogenesis or to be implicated in neurodegenerative disease, including the canonical Wnt/β-catenin and the Alzheimer’s disease/presenilin signaling pathways. Dysregulation of the Wnt/β-catenin pathway is confirmed by Western blot demonstration of cytosolic sequestration of β-catenin during in vitro differentiation of the SH-SY5Y cells toward the neuronal phenotype. We also demonstrate that two key transcription factor genes known to be regulated by Wnt signaling and to be vital for the generation and function of dopaminergic neurons; i.e., Lmx1a and Engrailed 1, are down-regulated in the HPRT knockdown SH-SY5Y cells. In addition to the Wnt signaling aberration, we found that expression of presenilin-1 shows a severely disturbed expression in HPRT-deficient cells, reflected by marked instability of the 23kDa C-terminal fragment of presenilin-1 in knockdown cells. Western blot analysis of primary cultures of two LND patients with 2.5% and 0% residual HPRT activity also shows dysregulated b-catenin and presenilin-1 expression, including elevated levels of cytosolic phospho-catenin and, in one of the two patient cells, failure of nuclear transport. Similarly, the presenilin-1 processing defect was most clearly demonstrated by markedly increased levels of both the N-terminal and C-terminal presenilin-1 fragments in the human cell line with no detectable residual enzyme activity but less marked over-expression in the cell with 2.5% residual enzyme activity. These demonstrations of dysregulated Wnt and presenilin-1 signaling and impaired expression of transcription factors necessary for dopaminergic development reveal broad pleitropic neuro-regulatory defects played by HPRT and suggest new directions for investigating mechanisms of aberrant neurogenesis and neuropathology in LND and potential new targets for restoration of effective signaling in this neuro-developmental defect. For microarray analysis, RNAs from triplicate independent cultures of vector-infected HPRT knockdown and control fibroblasts were prepared separately, pooled and used to prepare cRNA and finally subjected to microarray transcriptional analysis in triplicate. The integrity of total RNA from HPRT knockdown and control cells was confirmed by bioanalyzer (Agilent Technologies, Santa Clara, CA). The quality of total RNA samples from HPRT knockdown and control cells was assessed by 2100 bioanalyzer before application to microarray analysis. We determined that the RNA integrity number (RIN), (maximal degradation = 1; maximal molecular integrity = 10) was 10 for both the normal and knockdown cells (data not shown), indicating that isolated RNA samples were of sufficiently high quality to permit subsequent preparation of cDNA for microarray analysis. Microarray transcriptional analysis was performed in triplicate using the HumanWG-6 v3.0 Expression BeadChip system (Illumina, San Diego, CA). All reagents were obtained from HumanWG-6 v.3 Expression BeadChip Kit (Illumina) and all experimental processes were carried out according to manufacturer’s instruction (Illumina). After scanning of hybridized BeadChip, quantitation of slide images were performed using Illumina’s BeadArray software and the raw data were normalized by Loess normalization method, and then the normalized raw data in BeadStudio was exported to GeneSpring GX 10.0.2 (Agilent, Santa Clara, CA). For identification of genes significantly altered in knockdown cell compared with the control normal gene set, total detected entities were filtered by signal intensity value (upper cut-off 100th and lower cut-off 20th percentile) and error (coefficient of variation: CV < 50.0 percent) to remove very low signal entities and to select reproducible signal values of entities among the replicated experiments, respectively. In statistical analysis, t-test unpaired (p < 0.05) was applied and all significant changes above 2-fold were selected. Signals were selected if they were above microarray background (detection p-value < 0.05) in either all six experiments or in at least three knockdown or control experiments. Analysis of GO, GSEA and signaling pathway was carried out using GeneSpring GX 10.0.2 (Agilent) and the PANTHER Classification System (http://www.pantherdb.org/). In the analysis of signaling pathways using GeneSpring GX 10.0.2 (Agilent), a total of 140 cellular pathways were identified. For GSEA analysis, we used the false discovery rate (FDR) of <0.4 and p ≤ 0.1.
2010-09-25 | E-GEOD-24345 | biostudies-arrayexpress
Project description:ChIP-seq analysis of LSD2-depleted HepG2 cells revealed that many of the target genes were related to lipid metabolism. We found that LSD2 is an important epigenetic regulator of hepatic lipid metabolism. Examination of LSD2/H3K4me1 interaction in control and LSD2-knockdowned HepG2 cells.
2015-02-02 | E-GEOD-59830 | biostudies-arrayexpress
Project description:Globel gene expression was analyzed by RNA-seq to study the role of lincRNA TUNA in pluripotent mouse embryonic stem cells. mRNA profiles of CCE mES cells with shRNA-mediated depletion of lincRNA TUNA on days 2, 4, and 6, compared to a control shRNA.
2014-02-03 | E-GEOD-46730 | biostudies-arrayexpress
Project description:Hox genes are key regulators of development. In mammals, the study of these genes is greatly confounded by their large number, overlapping functions, and their interspersed shared enhancers. In this report, we describe a novel recombineering strategy that was used to introduce simultaneous frameshift mutations into the flanking Hoxa9, Hoxa10, and Hoxa11 genes, as well as their paralogs on the HoxD cluster. The resulting mutant mice displayed dramatic homeotic transformations of the reproductive tracts, with uterus anteriorized towards oviduct and the vas deferens anteriorized towards epididymis. The Hoxa9,10,11 mutant mice provided a sensitized genetic background that allowed the discovery of Hoxd9,10,11 reproductive tract patterning function. Both shared and distinct Hox functions were defined. The HoxD genes played a crucial role in the regulation of the uterine immune function. Non-coding nonpolyadenylated RNAs were among the key Hox targets. In addition we observed a surprising anti-dogmatic posteriorization of the uterine epithelium. Reproductive tracts were collected from WT and Hox mutant mice (n=3/genotype) aged 3-7 months in order to characterize the molecular changes caused by mutation of Hoxa9,10,11 and Hoxd9,10,11. Female mice were staged and collected in diestrus.
2013-08-01 | E-GEOD-42339 | biostudies-arrayexpress
Project description:We identified a novel recurrent genetic lesion in T-LGL. Mutations of the tumour suppressor gene TNFAIP3 causing amino-acid exchanges or protein truncations were seen in 3/39 cases (8%). RNA sequencing (Illumina HiSeq 2500) of 5 index patients with paired tumor and non-tumor samples.
2016-06-12 | E-GEOD-69735 | biostudies-arrayexpress
Project description:ChIP-seq analysis of HepG2 cells revealed that many of the target genes of LSD2 were related to lipid metabolism. We found that LSD2 is an important epigenetic regulator of hepatic lipid metabolism. Examination of LSD2/DNA interaction in HepG2 cells in normal condition.
2015-02-02 | E-GEOD-59694 | biostudies-arrayexpress
Project description:Comparing gene expression profile in 3T3-F442A adipocytes with shRNA against TRPV4 or GFP. TRPV4 is an ion channel expressed in adipocytes. Results provided information that how loss-of-function of TRPV4 affects gene expression in adipocytes. 4 samples were analyzed as two groups: shGFP (control) and shTRPV4 (experimental). Each group has two replicates.
2012-10-12 | E-GEOD-40280 | biostudies-arrayexpress
Project description:Increasing studies suggest that SALL4 may play vital roles in leukemogenesis. We have used chromatin immunoprecipitation followed by microarray hybridization as a screening tool to determine potential genes that may account for the role SALL4 plays in leukemogenesis. Analysis of SALL4 binding sites reveals that genes involved in cell death, cancer, DNA replication/repair, and cell cycle were highly enriched (p<0.05). These genes include 38 important apoptosis-inducing genes (TNF, TP53, PTEN, CARD9, CARD11, CYCS, LTA) and apoptosis-inhibiting genes (Bmi-1, BCL2, XIAP, DAD1, TEGT). Real-time PCR has shown that expression levels of these genes changed significantly after SALL4 knockdown, which ubiquitously led to cell apoptosis. Flow cytometry revealed that reduction of SALL4 expression in NB4 and other leukemia cell lines dramatically increased caspase-3, Annexin V, and DNA fragmentation activity. Bromodeoxyuridine-incorporation assays showed decreased numbers of S phase cells and increased numbers of G1- and G2- phase cells indicating reduced DNA synthesis, consistent with results from cell proliferation assays. In addition, NB4 cells that express low levels of SALL4 have significantly decreased tumorigenecity in immunodeficient mice. Our studies provide a foundation in the development of leukemia stem cell-specific therapy by targeting SALL4. Keywords: ChIP-chip The global targets of SALL4 were determined using a NimbleGen promoter tiling array (2.7kb, information below) in the human cell line NB4. Because SALL4 is thought to play a significant role in leukemogenesis we focused primarily on genes involved in apoptosis and only used the ChIP-chip array as a screening tool to identify potential genes that may bind these genes. Upon identification and verification of apoptosis genes bound by SALL4 we utilized functional assays to determine the effect of SALL4 on these genes. In addition, gene expression profiling was used to determine pathways functionally altered by Sall4 reduction. Source: UCSC Build: HG18 Probe Length: 50-75mer Median Probe Spacing: 100bp Probes per Array: 385,000 Feature Size: 16μm x 16μm Array Dimensions: 17.4mm x 13mm Overall Slide Dimensions: 1" x 3" (25 x 75mm) Recommended Storage: Store arrays desiccated at room temperature. Promoter tiling from 2200bp upstream and 500bp downstream of the transcription start site for RefSeq genes.
2008-05-17 | E-GEOD-10734 | biostudies-arrayexpress
Project description:Recent studies have shown that the RNA binding protein Musashi 2 (Msi2) plays prominent roles during development and leukemia. Additionally, in embryonic stem cells (ESC) undergoing the early stages of differentiation, Msi2 has been shown to associate with Sox2, which is required for the self-renewal of ESC. These findings led us to examine the effects of Msi2 on the behavior of ESC. Using an shRNA sequence that targets Msi2 and a scrambled shRNA sequence, we determined that knockdown of Msi2 disrupts the self-renewal of ESC and promotes their differentiation. Collectively, our findings argue that Msi2 is required to support the self-renewal and pluripotency of ESC. We used microarrays to better understand global changes in ESC gene expression following the knockdown of the RNA-binding protein Msi2 as compared to control ESC expressing a scrambled shRNA. Mouse embryonic stem cells (D3) were treated with lentivirus engineered for the expression of a Msi2 targeting shRNA sequence or scrambled (control) shRNA sequence. Following selection for infected cells with puromycin, cells were subcultured at low density and allowed to grow for 4 days (see treatment protocol) before RNA was extracted. RNA was collected and analyzed one time for each of the two samples.
2012-08-25 | E-GEOD-33882 | biostudies-arrayexpress