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GSE3188
Homo sapiens
9 Downloadable Samples
Affymetrix Human Genome U133A Array (hgu133a)
Description
The response of cells to hypoxia is characterised by co-ordinated regulation of many genes. Studies of the regulation of the expression of many of these genes by oxygen has implicated a role for the heterodimeric transcription factor hypoxia inducible factor (HIF). The mechanism of oxygen sensing which controls this heterodimeric factor is via oxygen dependent prolyl and asparaginyl hydroxylation by specific 2-oxoglutarate dependent dioxygenases (PHD1, PHD2, PHD3 and FIH-1). Whilst HIF appears to have a major role in hypoxic regulation of gene expression, it is unclear to what extent other transcriptional mechanisms are also involved in the response to hypoxia. The extent to which 2-oxoglutarate dependent dioxygenases are responsible for the oxygen sensing mechanism in HIF-independent hypoxic gene regulation is also unclear. Both the prolyl and asparaginyl hydroxylases can be inhibited by dimethyloxalylglycine (DMOG). Such inhibition can produce activation of the HIF system with enhanced transcription of target genes and might have a role in the therapy of ischaemic disease. We have examined the extent to which the HIF system contributes to the regulation of gene expression by hypoxia, to what extent 2-oxoglutarate dependent dioxygenase inhibitor can mimic the hypoxic response and the nature of the global transcriptional response to hypoxia. We have utilised microarray assays of mRNA abundance to examine the gene expression changes in response to hypoxia and to DMOG. We demonstrate a large number of hypoxically regulated genes, both known and novel, and find a surprisingly high level of mimicry of the hypoxic response by use of the 2-oxoglutarate dependent dioxygenase inhibitor, dimethyloxalylglycine. We have also used microarray analysis of cells treated with small interfering RNA (siRNA) targeting HIF-1alpha and HIF-2alpha to demonstrate the differing contributions of each transcription factor to the transcriptional response to hypoxia. Candidate transcripts were confirmed using an independent microarray platform and real-time PCR. The results emphasise the critical role of the HIF system in the hypoxic response, whilst indicating the dominance of HIF-1alpha and defining genes that only respond to HIF-2alpha.
Publication Title
Concordant regulation of gene expression by hypoxia and 2-oxoglutarate-dependent dioxygenase inhibition: the role of HIF-1alpha, HIF-2alpha, and other pathways.
Sample Metadata Fields
No sample metadata fields
View Samples- Mus musculus
- 67 Downloadable Samples
Illumina HiSeq 2000
Description
Mutations in the mitochondrial DNA (mtDNA) have been proposed to be essential for metabolic adaptation, and because metabolism is intrinsically associated with multiple disease states, including obesity, we hypothesized that changes in the mtDNA would significantly influence adiposity and gene expression in response to diet. To test these predictions we used Mitochondrial-Nuclear eXchange mice, which have nuclear and mitochondrial genomes that have been exchanged from different M. musculus strains. Overall design: Purpose: Mutations in the mitochondrial DNA (mtDNA) have been proposed to be essential for metabolic adaptation, and because metabolism is intrinsically associated with multiple disease states, including obesity, we hypothesized that changes in the mtDNA would significantly influence adiposity and gene expression in response to diet. To test these predictions we used Mitochondrial-Nuclear eXchange mice, which have nuclear and mitochondrial genomes that have been exchanged from different M. musculus strains. Methods: Wild type (C57BL6/J – C57n:C57mt and C3H/HeN - C3Hn:C3Hmt) and MNX (C57n:C3Hmt and C3Hn:C57mt) mouse were weaned with Chor diet and continued with Chow or changed to high-fat diet from 6 to 12-13 weeks of age. RNA samples were isolated from white adipose tissues collected from epididymal (eWAT) and inguinal (iWAT) fat, representing visceral and subcutaneous fat depots, respectively with RNeasy kit (Qiagen). Reverse transcribed cDNA libraries were sequenced with an Illumina HiSeq 2000. Read mapping was conducted with a proprietary algorithm by Expression Analysis (www.q2labsolutions.com), and read counts were used as input for differential expression analysis in DESeq2 version 1.10.1, using default settings. Results: Using an optimized data analysis workflow, we mapped about 20 million sequence reads per sample to the mouse genome (build mm9). Transcriptional changes were interrogated for 961 genes previously reported to be associated with fat metabolism and 29,209 genes representing the entire mouse transcriptome. These results show that the C57 mtDNA increased the number of DE genes in response to high fat diet in mice harboring the C3H nuclear genome (209% increase; C3Hn:C57mt versus C3Hn:C3Hmt, 165/79) and the C3H mtDNA decreased response in animals carrying the C57 nucleus (46% decrease; C57n:C3Hmt versus C57n:C57mt, 112/206) in eWAT (Figure 2B). Similarly, the high fat diet resulted in 25 and 231 DE genes in the C3Hn:C3Hmt and C3Hn:C57mt iWAT, respectively, and 344 and 143 DE genes in C57n:C57mt and C57n:C3Hmt iWAT. This corresponded to a 924% increase in the number of DE genes responding to high fat diet C3Hn:C57mt versus C3Hn:C3Hmt, and a decreased response (58% decrease) in C57n:C3Hmt relative to C57n:C57mt iWAT. Further analysis showed that each MNX and corresponding wild-type shared and had distinct DE genes in eWAT and iWAT. Conclusions: Results also show that the degree of transcriptional response influenced by the mtDNA can vary based upon the type of adipose tissue, suggesting that mtDNA background can have varying effects on the number of nuclear genes differentially responding to stimuli, depending upon tissue and location.
Publication Title
Mitochondrial - nuclear genetic interaction modulates whole body metabolism, adiposity and gene expression in vivo.
Sample Metadata Fields
Specimen part, Subject
View Samples- Homo sapiens
- 18 Downloadable Samples
- Illumina Genome Analyzer IIx
Description
Cells adapt to environmental changes, including fluctuations in oxygen levels, through the induction of specific gene expression programs. To identify genes regulated by hypoxia at the transcriptional level, we pulse-labeled HUVEC cells with 4-thiouridine and sequenced nascent transcripts. Then, we searched genome-wide binding profiles from the ENCODE project for factors that correlated with changes in transcription and identified binding of several components of the Sin3A co-repressor complex, including SIN3A, SAP30 and HDAC1/2, proximal to genes repressed by hypoxia. SIN3A interference revealed that it participates in the downregulation of 75% of the hypoxia-repressed genes in endothelial cells. Unexpectedly, it also blunted the induction of 47% of the upregulated genes, suggesting a role for this corepressor in gene induction. In agreement, ChIP-seq experiments showed that SIN3A preferentially localizes to the promoter region of actively transcribed genes and that SIN3A signal was enriched in hypoxia-repressed genes, prior exposure to the stimulus. Importantly, SINA3 occupancy was not altered by hypoxia in spite of changes in H3K27ac signal. In summary, our results reveal a prominent role for SIN3A in the transcriptional response to hypoxia and suggest a model where modulation of the associated histone deacetylase activity, rather than its recruitment, determines the transcriptional output. Overall design: Exponentially growing non-synchronized HUVEC were exposed to normoxia or hypoxia (21% or 1% oxygen respectively) for 8 hours and pulse-labelled with 4-thiouridine during the last two hours of treatment. RNA was extracted from samples in each condition (total RNA) and an aliquot was subjected to affinity chromatography to purify the 4-thiouridine-labelled (newly transcribed RNA, Newly Tr) and non-labelled (Pre-existent) RNA fractions. All three RNA fractions (total, newly transcribed and pre-existent) from each sample were analyzed by high-throughput sequencing. Submission includes 12 samples corresponding to 3 independent biological replicates.
Publication Title
The SIN3A histone deacetylase complex is required for a complete transcriptional response to hypoxia.
Sample Metadata Fields
Cell line, Treatment, Subject
View Samples- Homo sapiens
- 8 Downloadable Samples
- Illumina HiSeq 1500
Description
Cells adapt to environmental changes, including fluctuations in oxygen levels, through the induction of specific gene expression programs. To identify genes regulated by hypoxia at the transcriptional level, we pulse-labeled HUVEC cells with 4-thiouridine and sequenced nascent transcripts. Then, we searched genome-wide binding profiles from the ENCODE project for factors that correlated with changes in transcription and identified binding of several components of the Sin3A co-repressor complex, including SIN3A, SAP30 and HDAC1/2, proximal to genes repressed by hypoxia. SIN3A interference revealed that it participates in the downregulation of 75% of the hypoxia-repressed genes in endothelial cells. Unexpectedly, it also blunted the induction of 47% of the upregulated genes, suggesting a role for this corepressor in gene induction. In agreement, ChIP-seq experiments showed that SIN3A preferentially localizes to the promoter region of actively transcribed genes and that SIN3A signal was enriched in hypoxia-repressed genes, prior exposure to the stimulus. Importantly, SINA3 occupancy was not altered by hypoxia in spite of changes in H3K27ac signal. In summary, our results reveal a prominent role for SIN3A in the transcriptional response to hypoxia and suggest a model where modulation of the associated histone deacetylase activity, rather than its recruitment, determines the transcriptional output. Overall design: Exponentially growing non-synchronized HUVEC were transduced with lentiviral particles encoding for shRNA targeting EPAS1 or control shRNA. 72h after infection, cells were exposed to normoxia or hypoxia (21% or 1% oxygen respectively) for 8 hours and pulse-labelled with 4-thiouridine during the last two hours of treatment. RNA was extracted from samples in each condition (total RNA) and an aliquot subjected to affinity chromatography to purify the 4-thiouridine-labelled RNA fraction (newly transcribed RNA, Newly Tr). Both RNA fractions from each condition were analyzed by high-throughput sequencing. Data includes 8 samples from a single biological replicate.
Publication Title
The SIN3A histone deacetylase complex is required for a complete transcriptional response to hypoxia.
Sample Metadata Fields
Cell line, Subject
View Samples- Homo sapiens
- 24 Downloadable Samples
- Illumina HiSeq 4000
Description
Hypoxia inducible factor (HIF) is the major transcriptional regulator of cellular responses to hypoxia. The two principal HIF-a isoforms, HIF-1a and HIF-2a, are progressively stabilized in response to hypoxia and form heterodimers with HIF-1b to activate a broad range of transcriptional responses. Here we report on the pan-genomic distribution of isoform-specific HIF binding in response to hypoxia of varying severity and duration, and in response to genetic ablation of each HIF-a isoform. Our findings reveal that, despite an identical consensus recognition sequence in DNA, each HIF heterodimer loads progressively at a distinct repertoire of cell-type specific sites across the genome, with little evidence of redistribution under any of the conditions examined. Marked biases towards promoter proximal binding of HIF-1 and promoter distant binding of HIF-2 were observed under all conditions and were consistent in multiple cell type. The findings imply that each HIF isoform has an inherent property that determines its binding distribution across the genome, which might be exploited to therapeutically target the specific transcriptional output of each isoform independently. Overall design: RNA_seq analysis of hypoxic gene regulation in HKC8 and HepG2 cell lines and in RCC4 cell lines stably transfected with wtVHL
Publication Title
Hypoxia drives glucose transporter 3 expression through hypoxia-inducible transcription factor (HIF)-mediated induction of the long noncoding RNA NICI.
Sample Metadata Fields
Specimen part, Cell line, Subject
View Samples- Rattus norvegicus
- 8 Downloadable Samples
- Affymetrix Rat Genome 230 2.0 Array (rat2302)
Description
Silicon (Si) has long been known to play a major physiological role in certain organisms, including some sponges and many diatoms and higher plants, leading to the recent identification of multiple proteins responsible for silicon transport in a range of algal and plant species. In mammals, despite several convincing studies suggesting that silicon is an important factor in bone development and connective tissue health, there is a critical lack of understanding in biochemical pathways that enable silicon homeostasis. Here we report the identification of a mammalian efflux silicon transporter, namely Slc34a2 (also known as NaPiIIb), which was upregulated in the kidneys of rats following chronic dietary silicon deprivation. When heterologously expressed in Xenopus laevis oocytes, the protein displayed marked silicon transport activity, specifically efflux, comparable to plant OsLsi2 transfected in the same fashion and independent of sodium and/or phosphate influx. This is the first evidence for a specific active transporter protein for silicon in mammals and suggests an important role for silicon in vertebrates.
Publication Title
Identification of a mammalian silicon transporter.
Sample Metadata Fields
Sex, Age, Specimen part
View Samples- Homo sapiens
- 8 Downloadable Samples
- Affymetrix Human Genome U133 Plus 2.0 Array (hgu133plus2)
Description
High levels of oxidative stress and an associated neuronal DDR occur at the earliest stages of Alzheimer pathology (low Braak stage), and is associated with cognitive impairment.
Publication Title
Neuronal DNA damage response-associated dysregulation of signalling pathways and cholesterol metabolism at the earliest stages of Alzheimer-type pathology.
Sample Metadata Fields
Specimen part
View Samples- Mus musculus
- 6 Downloadable Samples
- Affymetrix Mouse Genome 430 2.0 Array (mouse4302)
Description
Neutrophils were isolated form peripheral blood of wildtype and Phd3 null mice, cultured for 4 hours in hypoxia (3% O2) and micro array analysis performed
Publication Title
Prolyl hydroxylase 3 (PHD3) is essential for hypoxic regulation of neutrophilic inflammation in humans and mice.
Sample Metadata Fields
Specimen part, Treatment
View Samples- Mus musculus
- 86 Downloadable Samples
- Affymetrix Mouse Genome 430 2.0 Array (mouse4302)
Description
This SuperSeries is composed of the SubSeries listed below.
Publication Title
Dynamic regulatory network controlling TH17 cell differentiation.
Sample Metadata Fields
Specimen part, Treatment
View Samples- Mus musculus
- 61 Downloadable Samples
- Illumina Genome Analyzer
Description
Despite their enormous importance, the molecular circuits that control the differentiation of Th17 cells remain largely unknown. Recent studies have reconstructed regulatory networks in mammalian cells, but have focused on short-term responses and relied on perturbation approaches that cannot be applied to primary T cells. Here, we develop a systematic strategy – combining transcriptional profiling at high temporal resolution, novel computational algorithms, and innovative nanowire-based tools for performing gene perturbations in primary T cells – to derive and experimentally validate a temporal model of the dynamic regulatory network that controls Th17 differentiation. The network is arranged into two self-reinforcing and mutually antagonistic modules that either suppress or promote Th17 differentiation. The two modules contain 12 novel regulators with no previous implication in Th17 differentiation, which may be essential to maintain the appropriate balance of Th17 and other CD4+ T cell subsets. Overall, our study identifies and validates 39 regulatory factors that are embedded within a comprehensive temporal network and identifies novel drug targets and organizational principles for the differentiation of Th17 cells. Overall design: RNA-seq of knockdown of 12 genes in Th17 cell differentiation
Publication Title
Dynamic regulatory network controlling TH17 cell differentiation.
Sample Metadata Fields
Specimen part, Cell line, Treatment, Subject
View Samples