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Revisiting Global Gene Expression Analysis (RNA-Seq)

ABSTRACT: Gene expression analysis is a widely used and powerful method for investigating the transcriptional behavior of biological systems, for classifying cell states in disease and for many other purposes. Recent studies indicate that common assumptions currently embedded in experimental and analytical practices can lead to misinterpretation of global gene expression data. We discuss these assumptions and describe solutions that should minimize erroneous interpretation of gene expression data from multiple analysis platforms. Polyadenylated RNA depleted of ribosomal content was used for preparation of two independent sequencing libraries (low-Myc & high-Myc). A panel of synthetic RNA's was added to these populations, based on cell number.

ORGANISM(S): Homo sapiens

SUBMITTER: Richard Young

PROVIDER: E-GEOD-40783 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Revisiting global gene expression analysis.

Lovén Jakob J Orlando David A DA Sigova Alla A AA Lin Charles Y CY Rahl Peter B PB Burge Christopher B CB Levens David L DL Lee Tong Ihn TI Young Richard A RA

Cell 20121001 3

Gene expression analysis is a widely used and powerful method for investigating the transcriptional behavior of biological systems, for classifying cell states in disease, and for many other purposes. Recent studies indicate that common assumptions currently embedded in experimental and analytical practices can lead to misinterpretation of global gene expression data. We discuss these assumptions and describe solutions that should minimize erroneous interpretation of gene expression data from mu ...[more]

PMID: 23101621

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Revisiting Global Gene Expression Analysis (Microarray)

Project description:Gene expression analysis is a widely used and powerful method for investigating the transcriptional behavior of biological systems, for classifying cell states in disease and for many other purposes. Recent studies indicate that common assumptions currently embedded in experimental and analytical practices can lead to misinterpretation of global gene expression data. We discuss these assumptions and describe solutions that should minimize erroneous interpretation of gene expression data from multiple analysis platforms. RNA was extracted from cells expressing low or high levels of c-Myc. A panel of synthetic RNA's was added to these populations, based on cell number. Total RNA was then extracted and processed on Affymetrix microarrays in biological duplicate.

2012-10-25 | E-GEOD-40782 | biostudies-arrayexpress

Chromatin occupancy of key transcriptional regulators in P493-6 cells with various levels of c-Myc expression

Project description:P493-6 cells are immortalized human peripheral B cells that carry a conditional, tetracycline-regulated myc gene. We present ChIP-seq analysis of key transcritional regulators in P493-6 cells expressing various levels of c-Myc: 0hr (low c-Myc levels), 1hr (intermediate c-Myc levels), 24hr (very high c-Myc levels) and No Tet (steady-state c-Myc levels). Brd4, c-Myc, Max, Med1, RNAPII, and the chromatin modification H3K27Ac were profiled in P493-6 cells

2012-11-14 | E-GEOD-42262 | biostudies-arrayexpress

Selective transcriptional regulation by Myc in cellular growth control and lymphomagenesis [ChIP-Seq]

Project description:The c-myc proto-oncogene product, Myc, is a transcription factor that binds thousands of genomic loci. Recent work suggested that rather than up- and down-regulating selected groups of genes, Myc targets all active promoters and enhancers in the genome (a phenomenon termed "invasion") and acts as a general amplifier of transcription. However, the available data did not readily discriminate between direct and indirect effects of Myc on RNA biogenesis. We addressed this issue with genome-wide chromatin immunoprecipitation and RNA expression profiles during B-cell lymphomagenesis in mice, in cultured B-cells and fibroblasts. Consistent with long-standing observations, we detected general increases in total RNA or mRNA copies per cell (hereby termed "amplification") when comparing actively proliferating cells with control quiescent cells: this was true whether cells were stimulated by mitogens (requiring endogenous Myc for a proliferative response) or by deregulated, oncogenic Myc activity. RNA amplification and promoter/enhancer invasion by Myc were separable phenomena that could occur without one another. Moreover, whether or not associated with RNA amplification, Myc drove the differential expression of distinct subsets of target genes. Hence, while having the potential to interact with all active/poised regulatory elements in the genome, Myc does not directly act as a global transcriptional amplifier. Instead, our results imply that Myc activates and represses transcription of discrete gene sets, leading to changes in cellular state that can in turn feed back on global RNA production and turnover. Mapping c-Myc, RNAPII and H3K4me3, H3K4me1 and H3K27ac in four different models of Myc regulation Note: GEO Sample GSM894093 was used as input sample to generate processed data files P493.Myc.NoMyc.bed, P493.Myc.LowMyc.bed, P493.Myc.HighMyc.bed, P493.Myc.t1h.bed, P493.Myc.t24h.bed, P493.Pol2.NoMyc.bed, P493.Pol2.t24h.bed

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2012-04-04 | E-GEOD-36354 | biostudies-arrayexpress

Selective transcriptional regulation by Myc in cellular growth control and lymphomagenesis

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2014-07-07 | E-GEOD-51011 | biostudies-arrayexpress

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Project description:This SuperSeries is composed of the SubSeries listed below. Refer to individual Series

2015-02-01 | E-GEOD-59480 | biostudies-arrayexpress

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Project description:This SuperSeries is composed of the SubSeries listed below. Refer to individual Series

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Gene expression analysis in cytoplasmic and nuclear fractions of P493-6 B cells with low c-myc expression to assess long noncoding RNA localization

Project description:Myc is a well known transcription factor with important roles in cell cycle, apoptosis and cellular transformation. Long non-coding (lnc)RNAs have recently emerged as a important class of regulatory RNAs. Here, we show that lncRNAs are an extensive component of the Myc-regulated transcriptional program. Using the P493-6 inducible Myc model we demonstrate that both Myc-induced mRNAs and lncRNAs were significant enriched for Myc binding sites. In contrast to Myc-repressed mRNAs, Myc-repressed lncRNAs were significantly enriched for Myc binding sites. Subcellular localization analysis revealed that Myc-repressed lncRNAs and mRNAs are enriched in the nucleus while Myc-induced lncRNAs and mRNAs are enriched both in the cytoplasm and nucleus. Parallel analysis of differentially expressed lncRNAs and mRNAs identified 105 lncRNA-mRNA pairs that were in close vicinity, indicative for regulation in cis. To support the potential relevance of the Myc-regulated lncRNAs in cellular transformation, we analyzed their expression in primary Myc-high and Myc-low B-cell lymphomas. In total, 54% of the lncRNAs differentially expressed between the lymphoma subsets were identified as Myc-regulated in P493-6 cells. This study is the first to show that lncRNAs are an important factor within the Myc-regulated transcriptional program and indicates a marked difference between Myc-repressed lncRNAs and mRNAs. Arrays: The total, cy3 labeled fraction and the cytoplasmic or nuclear, cy5 labeled fraction was hybridized on the same array. Data was imported and analyzed as a single color array. Expression of all known mRNAs and >10 000 lncRNAs was assessed in P493-6 B cells with low c-myc levels

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