Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Transcription profiling of mouse ES cells: WT, HIF-1KO, HIF-2KO cells treated under 21% or 1.5% oxygen to indentify hypoxia induced genes

ABSTRACT: WT, HIF-1KO, HIF-2KO ES cells were treated under 21% or 1.5% oxygen. HIF-1 target genes will be identified in HIF-2KO ES cell while HIF-2 target genes will be identified in HIF-1KO ES cells. WT ES cells expressing both HIF-1 and HIF-2 protein will be useful to identify hypoxia indicible genes induced by HIF-1 and HIF-2.

ORGANISM(S): Mus musculus

SUBMITTER: Sangeeta Iyer

PROVIDER: E-MEXP-392 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

Differential regulation of the transcriptional activities of hypoxia-inducible factor 1 alpha (HIF-1alpha) and HIF-2alpha in stem cells.

Hu Cheng-Jun CJ Iyer Sangeeta S Sataur Aneesa A Covello Kelly L KL Chodosh Lewis A LA Simon M Celeste MC

Molecular and cellular biology 20060501 9

Transcriptional responses to hypoxia are primarily mediated by hypoxia-inducible factors (HIFs), HIF-1alpha and HIF-2alpha. The HIF-1alpha and HIF-2alpha subunits are structurally similar in their DNA binding and dimerization domains but differ in their transactivation domains, implying they may have unique target genes and require distinct transcriptional cofactors. Our previous results demonstrated that HIF-1alpha and HIF-2alpha regulate distinct target genes. Here, we report that HIF-2alpha i ...[more]

PMID: 16611993

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Project description:Low-oxygen tolerance is supported by an adaptive response that includes a coordinate shift in metabolism and the activation of a transcriptional program that is driven by the hypoxia-inducible factor (HIF) pathway. The precise contribution of HIF-1 in the adaptive response, however, has not been determined. Here we investigate how HIF-1 influences hypoxic adaptation throughout Drosophila development. We find that hypoxic-induced transcriptional changes are comprised of HIF-dependent and HIF-independent pathways that are distinct and separable. We show that normoxic set-points of carbohydrate metabolites are significantly altered in dHIF mutants and that these animals are unable to mobilize glycogen in hypoxia. Furthermore, we find that the estrogen-related receptor (dERR), which is a global regulator of aerobic glycolysis in larvae, is required for a competent hypoxic response. dERR binds to dHIF and participates in the HIF-dependent transcriptional program in hypoxia. In addition, dERR acts in the absence of dHIF in hypoxia and a significant portion of HIF-independent transcriptional responses can be attributed to dERR actions, including upregulation of glycolytic transcripts. These results indicate that competent hypoxic responses arise from complex interactions between HIF-dependent and -independent mechanisms, and that dERR plays a central role in both of these programs. Fly eggs were collected onto egg caps with yeast paste . The caps were replaced after overnight incubation periods and kept at 25M-BM-:C, until mid-L2. At mid-L2, larvae were transferred to a fresh egg cap with blue yeast paste (0.3% bromophenol blue), and allowed to develop until achieving the partial clear gut L3 stage (-10 to -4 hrs prior to metamorphic onset). Appropriately staged animals were moved to fresh agar plates and allowed to age an additional 6 hours at 25M-BM-:C (normoxic treatment). Alternatively, animals were placed in an airtight Modular Incubator Chamber for 6 hours at 25M-BM-:C after a gas mixture containing 4% oxygen balanced with nitrogen was flashed into the chamber (hypoxic treatment). Mutant larvae were sorted for the absence of GFP expression using dissecting stereoscope with fluorescence at mid-L2. Microarray analyses were performed on at least three biological replicates of w1118, dHIF mutants, or dERR mutants at the partial clear gut third instar larval stage and were treated for 6 hours in normoxia or 4% O2. For each biological replicate, at least 10 larvae were collected and washed with 1M-CM-^WPBS. Larvae were placed in TRIzol (Invitrogen, Carlsbad, CA) and homogenized using VWR disposable pellet mixer. Total RNA was isolated using a TRIzol/RQ1 DNase (Promega, Madison, WI) hybrid extraction protocol. Template labelings were performed using the GeneChip 3' IVT Express Kit according to the manufacturerM-bM-^@M-^Ys specifications (Affymetrix, Santa Clara, CA). Hybridizations to Affymetrix GeneChip Drosophila Genome 2.0 arrays were performed using the manufacturers recommendations. Every chip was scanned at a high resolution by the Affymetrix GeneChipM-BM-. Scanner 3000 according to the GeneChip Expression Analysis Technical Manual procedures (Affymetrix, Santa Clara, CA).

Dataset Information

Transcription profiling of mouse ES cells: WT, HIF-1KO, HIF-2KO cells treated under 21% or 1.5% oxygen to indentify hypoxia induced genes

Publications

Differential regulation of the transcriptional activities of hypoxia-inducible factor 1 alpha (HIF-1alpha) and HIF-2alpha in stem cells.

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