The recently released Affymetrix Human Gene 1.0 ST array has two major differences compared with standard 3' based arrays: (1) it interrogates the entire mRNA transcript, and (2) it uses cDNA targets. To assess the impact of these differences on array performance, we performed series of comparative hybridizations between the Human Gene 1.0 ST and the Affymetrix HG-U133 Plus 2.0 and the Illumina HumanRef-8 BeadChip arrays. Additionally, both cRNA and cDNA targets were probed on the HG-U133 Plus 2.0 array. The results show that the overall reproducibility is best using the Gene 1.0 ST array. When looking only at the high intensity probes, the reproducibility of the Gene 1.0 ST array and the Illumina BeadChip array is equally good. Concordance of array results was assessed using different inter-platform mappings. The Gene 1.0 ST is most concordant with the HG-U133 array hybridized with cDNA targets, thus showing the impact of the target type. Agreements are better between platforms with designs which choose probes from the 3' end of the gene. Overall, the high degree of correspondence provides strong evidence for the reliability of the Gene 1.0 ST array.
Affymetrix Whole-Transcript Human Gene 1.0 ST array is highly concordant with standard 3' expression arrays.
No sample metadata fields
View SamplesRegulation of mRNA stability by RNA-protein interactions contributes significantly to quantitative aspects of gene expression. We have identified potential mRNA targets of the AU-rich element binding protein AUF1. Myc-tagged AUF1 p42 was induced in mouse NIH-3T3 cells and RNA-protein complexes isolated using anti-myc tag antibody beads. Bound mRNAs were analyzed with Affymetrix microarrays. We have identified 508 potential target mRNAs that were at least 3-fold enriched compared to control cells without myc-AUF1. 22.3% of the enriched mRNAs had an AU-rich cluster in the ARED Organism database, against 16.3% of non-enriched control mRNAs. The enrichment towards AU-rich elements was also visible by AREScore with an average value of 5.2 in the enriched mRNAs versus 4.2 in the control group. Yet, many mRNAs were enriched without a high ARE score suggesting that AUF1 has a broader binding spectrum than standard AUUUA repeats. AUF1 did not preferentially bind to unstable mRNAs. Still, some enriched mRNAs were highly unstable, as those of TNFSF11 (known as RANKL), KLF10, HES1, CCNT2, SMAD6, and BCL6. We have mapped some of the instability determinants. HES1 mRNA appeared to have a coding region determinant. Detailed analysis of the RANKL and BCL6 3UTR revealed for both that full instability required two elements, which are conserved in evolution. In RANKL mRNA both elements are AU-rich and separated by 30 bases, while in BCL6 mRNA one is AU-rich and 60 bases from a non AU-rich element that potentially forms a stem-loop structure.
Short-lived AUF1 p42-binding mRNAs of RANKL and BCL6 have two distinct instability elements each.
Cell line
View SamplesIn this study, we analyzed global liver gene expression in MICU1 knock-down (KD) mice. To generate liver-specific MICU1 KD mice, MICU1loxp/loxp male mice were treated with an AAV8-Cre under the control of a hepatocyte specific promoter (TBG). AAV8-TBG-Null treated littermates were used as controls. Liver samples were collected 3-5 weeks after injection. Knockdown was verified by protein and mRNA (94%, 98%, respectively). Mouse Gene 2.0 ST (Affymetrix, Santa Clara, CA) arrays were used to obtain global gene expression data.
MICU1 regulation of mitochondrial Ca(2+) uptake dictates survival and tissue regeneration.
Treatment
View SamplesThe discovery of mammalian cardiac progenitor cells has suggested that the heart consists of not only terminally differentiated beating cardiomyocytes, but also a population of self-renewing stem cells with the potential to generate new cardiomyocytes (Anderson, Self et al. 2007; Bearzi, Rota et al. 2007; Wu, Chien et al. 2008). A consequence of longevity is continual exposure to environmental and xenobiotic stresses, and recent literature suggests that hematopoietic stem cell pools tightly control cell health through upregulation of the integrated stress response and consequent cellular mechanisms such as apoptosis. However, whether or not this biological response is conserved in progenitor cells for later lineages of tissue specific stem cells is not well understood. Using human induced pluripotent stem cells (iPSC) of both cardiac progenitor and mature cardiomyocyte lineages, we found that the integrated stress response was upregulated in the iPSC cardiac progenitors leading to an increased sensitivity for apoptosis relative to the mature cardiomyocytes. Of interest, C/EBP homologous protein (CHOP) signaling plays a mechanistic role in the cell death phenotype observed in iPSC progenitors, by which depletion of CHOP prevents cell death following cellular stress by thapsigargin exposure. Our studies suggest that the integrated stress response plays a unique role in maintaining iPSC cardiac progenitor cellular integrity by removing unhealthy cells via apoptosis following environmental and xenobiotic stresses, thus preventing differentiation and self-renewal of damaged cells.
The Integrated Stress Response Regulates Cell Health of Cardiac Progenitors.
Specimen part, Treatment
View SamplesExpression profiling of Ewing sarcoma samples in the frame of the CIT program from the french Ligue Nationale Contre le Cancer (http://cit.ligue-cancer.net).
Common variants near TARDBP and EGR2 are associated with susceptibility to Ewing sarcoma.
Sex, Age
View SamplesTesticular gene expression changes with loss of Topaz1
TOPAZ1, a germ cell specific factor, is essential for male meiotic progression.
Specimen part
View SamplesRegeneration of lung epithelium is vital for maintaining airway function and integrity. An imbalance between epithelial damage and repair is at the basis of numerous chronic lung diseases such as asthma, COPD, pulmonary fibrosis and lung cancer. IGF (Insulin-like Growth Factors) signaling has been associated with most of these respiratory pathologies, although their mechanisms of action in this tissue remain poorly understood. Expression profiles analyses of IGF system genes performed in mouse lung support their functional implication in pulmonary ontogeny. Immuno-localization revealed high expression levels of Igf1r (Insulin-like Growth Factor 1 Receptor) in lung epithelial cells, alveolar macrophages and smooth muscle. To further understand the role of Igf1r in pulmonary homeostasis, two distinct lung epithelial-specific Igf1r mutant mice were generated and studied. The lack of Igf1r disturbed airway epithelial differentiation in adult mice revealed enhanced proliferation and altered morphology in distal airway club cells. During recovery after naphthalene-induced club cell injury, the kinetics of terminal bronchiolar epithelium regeneration was hindered in Igf1r mutants, revealing increased proliferation and delayed differentiation of club and ciliated cells. Amid airway restoration, lungs of Igf1r deficient mice showed increased levels of Igf1, Insr, Igfbp3 and epithelial precursor markers, reduced amounts of Scgb1a1 protein, and alterations in IGF signaling mediators. These results support the role of Igf1r in controlling the kinetics of cell proliferation and differentiation during pulmonary airway epithelial regeneration after injury. Overall design: Lung mRNA profiles of 3 months-old Igf1rfl/fl normal/control transgenic mice were generated by deep sequencing using Illumina GAIIx. ------------------------------------------- Submitter states "we use data on the absolute transcription levels (FPKM) of same IGF system genes on the adult "normal" mouse lung to compare them with those reported in the human adult lung (expressed in both as FPKM) (http://www.proteinatlas.org/)".
Involvement of Igf1r in Bronchiolar Epithelial Regeneration: Role during Repair Kinetics after Selective Club Cell Ablation.
Specimen part, Cell line, Subject
View SamplesMicroRNAs (miRNAs) are short noncoding RNA molecules regulating the expression of mRNAs. Target identification of miRNAs is computationally difficult due to the relatively low homology between miRNAs and their targets. We present here an experimental approach to target identification where the cartilage-specific miR-140 was overexpressed and silenced in cells it is normally expressed in separate experiments. Expression of mRNAs was profiled in both experiments and the intersection of mRNAs repressed by miR-140 overexpression and derepressed by silencing of miR-140 was identified. The intersection contained only 49 genes, although both treatments affected the accumulation of hundreds of mRNAs. These 49 genes showed a very strong enrichment for the miR-140 seed sequence implying that the approach is efficient and specific. 21 of these 49 genes were predicted to be direct targets based on the presence of the seed sequence. Interestingly, none of these were predicted by the published target prediction methods we used. One of the potential target mRNAs, Cxcl12, was experimentally validated by Northern blot analysis and a luciferase reporter assay.
Experimental identification of microRNA-140 targets by silencing and overexpressing miR-140.
No sample metadata fields
View SamplesInsight into the role of Insulin-like Growth Factor (IGF) in development of lungs has come from the study of genetically modified mice. IGF1 is a key factor during lung development. IGF1 deficiency in the neonatal mouse causes respiratory failure collapsed alveoli and altered alveolar septa. To further characterize IGF1 function during lung development we analyzed Igf1-/- mouse prenatal lungs in a C57Bl/6 genetic background. Mutant lungs showed disproportional hypoplasia, disorganized extracellular matrix and dilated alveolar capillaries. IGF1 target genes during lung maturation were identified by analyzing RNA differential expression in Igf1-/- lungs using microarrays.
Transcriptome analysis in prenatal IGF1-deficient mice identifies molecular pathways and target genes involved in distal lung differentiation.
Specimen part
View SamplesComparison of expression profile of Ewing''s sarcoma with cell of origin, mesenchymal stem cells with the goal of identifying novel therapeutic targets. Overall design: 3 Ewing''s cell lines compared to 2 MSC cell lines
Exploring the surfaceome of Ewing sarcoma identifies a new and unique therapeutic target.
No sample metadata fields
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