Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Immediate-Early and Delayed Primary Response Genes are Distinct in Function and Genomic Architecture

ABSTRACT: Transcriptional profiling time course of serum deprived quiescent human T98G cells follwing platelet derived growth factor (PDGF) stimulation in the absence and presence of the translation inhibitor, cycloheximide. RNA was harvested at 0.5, 2, and 4 hr following PDGF treatment. Keywords: time course Two-color microarrays with dye-swap; 3 biological replicates per condition, independently grown and harvested. One replicate per array.

ORGANISM(S): Homo sapiens

SUBMITTER: Steven Mullenbrock

PROVIDER: E-GEOD-8315 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

Immediate-early and delayed primary response genes are distinct in function and genomic architecture.

Tullai John W JW Schaffer Michael E ME Mullenbrock Steven S Sholder Gabriel G Kasif Simon S Cooper Geoffrey M GM

The Journal of biological chemistry 20070615 33

The transcriptional program induced by growth factor stimulation is classically described in two stages as follows: the rapid protein synthesis-independent induction of immediate-early genes, followed by the subsequent protein synthesis-dependent induction of secondary response genes. In this study, we obtained a comprehensive view of this transcriptional program. As expected, we identified both rapid and delayed gene inductions. Surprisingly, however, a large fraction of genes induced with dela ...[more]

PMID: 17575275

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Project description:Persistence of memory CD4+ T cells in ECs was coupled with the inactivation of FOXO3a transcriptional activities, which we have previously identified as a critical regulator of TCM survival. Indeed, expression levels of transcriptional targets of FOXO3a, endowed with pro-apoptotic and anti-proliferative functions, were lower in TCM and TEM from ECs as compared to ST individuals. Silencing the transcriptionally active form of FOXO3a by siRNA rescued TCM and TEM of STs from Fas-mediated apoptosis. Moreover the expression of FOXO3a dominant negative form (FOXO3a Nt) rescued the long-term survival of TCM from STs as these cells persisted as long as those derived from ECs. Overall, these studies indicate that FOXO3a activation is an important mediator of the shortened survival and heighteined turnover of TCM and TEM in chronic HIV infection. Targeting this pathway may provide a strategy to preserve memory T cell numbers in HIV infection. Keywords: comparative gene profile, cell-type comparison Isolation of CD4+T cell sub-populations. Peripheral blood mononuclear cells (PBMCs) from healthy adult individuals were isolated by Ficoll-HyPaque (Pharmacia) density gradient. We first enriched for CD4+ T cells using negative immunomagnetic beads selection (Automacs, Myltenii), cells were then labeled with anti-CD4-APCcy7, anti-CD45RA-ECD, anti-CD27-FITC and anti-CCR7-PEcy7 and sorted into Naive cells described as CD4+, CD45RA+, CD27+ and CCR7+, Central Memory cells (TCM) described as CD4+, CD45RA-, CD27+ and CCR7+ cells and Effector Memory cells (TEM) described as CD4+, CD45RA-, CD27- and CCR7- cells. Sorting was performed using a BDAria (BD Pharmingen). Purity of the TCM and TEM sub-populations was ranging from 96 to 99%. All procedures were done at 4C to avoid any changes in cell phenotype or gene expression. Sample RNA was extracted using an RNA extraction kit (Quiagen), then amplified using the MessageAmp RNA kit (Ambion) as per the manufacturer’s instructions. The amplified RNA (aRNA) was then verified for quality and quantity using the Agilent Bioanalyser and measuring the OD. Universal human RNA (Stratagene) was also prepared in the same way. Sample probes were prepared by direct labelling with 3 µg of the aRNA Cy-5 (R values) fluorescent dye while the universal RNA probes were prepared by direct labelling of universal aRNA with Cy-3 (G values). All patient samples were hybridized against amplified universal RNA at 37 ºC for 18h on a custom human Immune array. Detailed information on the labeling and hybridization procedures can be obtained at the Microarray Centre web page (University Health Network).

Dataset Information

Immediate-Early and Delayed Primary Response Genes are Distinct in Function and Genomic Architecture

Publications

Immediate-early and delayed primary response genes are distinct in function and genomic architecture.

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