Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Transcription profiling of human hGRbeta in the absence of hGR alpha

ABSTRACT: The human glucocorticoid receptor (GR) is expressed as two alternately spliced Cterminal isoforms: Î± and Î². In contrast to the canonical hGRÎ±, hGRÎ² is constitutively nuclear, is not thought to bind ligand, and is believed to affect gene transcription only by acting as a dominant negative to hGRÎ±. Microarray analysis indicated that hGRÎ², expressed in the absence of hGRÎ±, can regulate gene expression, and furthermore, occupation of hGRÎ² with the antagonist RU-486 diminishes that capacity. Experiment Overall Design: RNA preparation: Total RNA was extracted from 5x10 6 U-OFF or U-2 Experiment Overall Design: OSÎ² cells treated with either ethanol vehicle or 1Î¼M RU-486 for 6 hours using the RNAqueous Total RNA Isolation Kit (Ambion Inc. Austin, TX) according to manufacturerâs instructions. RNA was treated with DNase using the DNA-Free DNase Treatment and Removal Reagents (Ambion Inc.) according to manufacturerâs instructions prior to use with the microarray. Four pairs of RNA (vehicle vs. RU-486 treated) were harvested for each cell type to yield four biological replicates for gene expression analysis. Experiment Overall Design: Linear Amplification Label Protocol and Feature Extraction: Gene expression analysis was conducted using Agilent Human1Av2 arrays (Agilent Technologies, Palo Alto, CA). Total RNA was amplified using the Agilent Low RNA Input Fluorescent Linear Amplification Kit protocol. Starting with 500ng of total RNA, Cy3 or Cy5 labeled cRNA was produced according to manufacturerâs protocol. For each two color comparison, 750ng of each Cy3 and Cy5 labeled cRNAs were mixed and fragmented using the Agilent In Situ Hybridization Kit protocol. Hybridizations were performed for 17 hours in a rotating hybridization oven using the Agilent 60-mer oligo microarray processing protocol. Slides were washed as indicated in this protocol Experiment Overall Design: and then scanned with an Agilent Scanner. Data was obtained using the Agilent Feature Extraction software (v7.5), using defaults for all parameters.

ORGANISM(S): Homo sapiens

SUBMITTER: NIEHS Microarray Group

PROVIDER: E-GEOD-5310 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

Human glucocorticoid receptor beta binds RU-486 and is transcriptionally active.

Lewis-Tuffin Laura J LJ Jewell Christine M CM Bienstock Rachelle J RJ Collins Jennifer B JB Cidlowski John A JA

Molecular and cellular biology 20070122 6

Human glucocorticoid receptor (hGR) is expressed as two alternately spliced C-terminal isoforms, alpha and beta. In contrast to the canonical hGRalpha, hGRbeta is a nucleus-localized orphan receptor thought not to bind ligand and not to affect gene transcription other than by acting as a dominant negative to hGRalpha. Here we used confocal microscopy to examine the cellular localization of transiently expressed fluorescent protein-tagged hGRbeta in COS-1 and U-2 OS cells. Surprisingly, yellow fl ...[more]

PMID: 17242213

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Dataset Information

Transcription profiling of human hGRbeta in the absence of hGR alpha

Publications

Human glucocorticoid receptor beta binds RU-486 and is transcriptionally active.

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