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The p53 Target Gene Siva Enables Tumorigenesis

ABSTRACT: Siva is a p53target gene essential for p53-dependent apoptosis, although it can also promote proliferation through inhibition of p53 in some settings. Thus, the role of Siva in tumorigenesis remains unclear. Surprisingly, we find that Siva loss inhibits non-small cell lung cancer (NSCLC) development, suggesting that Siva facilitates tumorigenesis. Similarly, Siva knockdown in mouse and human NSCLC cell lines decreases proliferation and transformation. To gain an understanding of the molecular basis for the reduced proliferation and transformation resulting from Siva knockdown, we performed gene expression profiling experiments on LSZ4 and LSZ2 cells after Siva knockdown by lentiviral transduction with each of the two previously characterized shRNAs (shSiva1 and shSiva2) and each of two control shRNAs (shLacZ and shGFP). Hierarchical clustering revealed a notable difference in gene expression profiles between Siva knockdown and control samples. Differentially expressed genes were analyzed for enrichment in Gene Ontology (GO) terms using GeneSpring Analysis. We found that the most enriched GO terms included immune regulation – consistent with Siva’s known role in regulating NFкB signaling – as well as cell migration and metabolic processes. RNA from LSZ2 and LSZ4 non small cell lung cancer cells expressing control (shGFP1, shLacz) or Siva (shSiva1, shSiva2) hairpins was used. A total of seven control samples and eight test samples using different cell lines and hairpins were used.

ORGANISM(S): Mus musculus

SUBMITTER: Stephano Spano Mello

PROVIDER: E-GEOD-66126 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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The p53 Target Gene Siva Enables Tumorigenesis

Project description:Siva is a p53target gene essential for p53-dependent apoptosis, although it can also promote proliferation through inhibition of p53 in some settings. Thus, the role of Siva in tumorigenesis remains unclear. Surprisingly, we find that Siva loss inhibits non-small cell lung cancer (NSCLC) development, suggesting that Siva facilitates tumorigenesis. Similarly, Siva knockdown in mouse and human NSCLC cell lines decreases proliferation and transformation. To gain an understanding of the molecular basis for the reduced proliferation and transformation resulting from Siva knockdown, we performed gene expression profiling experiments on LSZ4 and LSZ2 cells after Siva knockdown by lentiviral transduction with each of the two previously characterized shRNAs (shSiva1 and shSiva2) and each of two control shRNAs (shLacZ and shGFP). Hierarchical clustering revealed a notable difference in gene expression profiles between Siva knockdown and control samples. Differentially expressed genes were analyzed for enrichment in Gene Ontology (GO) terms using GeneSpring Analysis. We found that the most enriched GO terms included immune regulation – consistent with Siva’s known role in regulating NFкB signaling – as well as cell migration and metabolic processes.

2015-05-15 | GSE66126 | GEO

Gene expression profiles upon knocking down Cops2 or Cops5 in mouse embryonic stem cells

Project description:To understand the role of Cops2 in mouse embryonic stem cells, we applied expression microarray analysis with GFP knockdown, Cops2 knockdown (2 different hairpins), and Cops5 knockdown mouse ESCs. Mouse ESCs were transfected with shGFP, shCops2(two different shRNAs), or shCops5 plasmids. 72 hours after transfection, cells were harvested for RNA prep and microarray analysis.

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Murine Myoblast C2C12 Cells: Control vs. Mybbp1a Knockdown

Project description:Transcriptional profiling of mouse myoblast cells comparing control vs. Mybbp1a knockdown. Stable clones of C2C12 cells harboring control or Mybbp1a-targeting shRNA were established and further pooled for analysis. Goal was to determine, based on the effects of Mybbp1a depletion on global gene expression, candidate downstream target genes of Mybbp1a, a putative transcriptional co-repressor. Two-condition experiment, control vs. Mybbp1a knockdown C2C12 cells (mixed stable clones). Biological replicates: 2.

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Comparison of gene expression in control and BRG1-knockdown H358 cell lines

Project description:Transcription was assessed in the human H358 NSCLC cell line after knockdown of BRG1, one of the mutually exclusive subunits of the SWI/SNF chromatin remodeling complex, by 2 different shRNAs.

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Gene expression profiles of RPMI8226 cells after knockdown of KDM3A and KLF2

Project description:Recent studies have implicated KDM3A, which catalyzes removal of H3K9 methylation, is associated with tumorigenesis. However, the biological role of KDM3A in multiple myeloma, has not been delineated. Here we identify KDM3A-KLF2-IRF4 axis dependence in multiple myeloma. We demonstrate that knockdown of KDM3A leads to apoptosis and significant growth inhibition in myeloma cells. Mechanistically, KDM3A directly regulates myeloma cell survival factor IRF4 expression through H3K9 demethylation at its promoter. We further show that KDM3A directly regulates KLF2 expression and that knockdown of KLF2 leads to growth inhibition in myeloma cells. The goal of this analysis is to identify genes whose expression changes after shRNA-mediated knockdown of KDM3A and KLF2 using the human U133 plus 2.0 Affymetrix GeneChip in myeloma cell line (RPMI8226). Two independent experiments were performed: 1. Myeloma cell line (RPMI8226) was transduced with either shRNAs targeting KDM3A (duplicate hairpins) or luciferase (control) in duplicate. The gene expression profiles of KDM3A knockdown cells were compared with that of control cells. A total of 6 RNA samples (4 KDM3A knockdown and 2 control) were analyzed. 2. Myeloma cell line (RPMI8226) was transduced with either shRNAs targeting KLF2 (duplicate hairpins) or luciferase (control) in duplicate. The gene expression profiles of KLF2 knockdown cells were compared with that of control cells. A total of 6 RNA samples (4 KLF2 knockdown and 2 control) were analyzed.

2015-02-18 | E-GEOD-55667 | biostudies-arrayexpress

Comparison of gene expression in control and BRG1-knockdown H358 cell lines

2014-09-23 | E-GEOD-58542 | biostudies-arrayexpress

Change in gene expression in Chlamydomonas reinhardtii upon heat shock

Project description:This was an accompanying experiment. Previously, the expression of genes hsp70A and hsp70B had been established as being tetrapyrrole-dependent (Kropat et al., 1997, 2000). Since both are well-known heat-shock proteins, we performed a one-step heat shock treatment by shifting the temperatures from 23M-BM-0C to 42M-BM-0C for 45 min. Moreover, this experiment served as an additional global control to further test the reliability of the microarray and our general conditions since the heat shock response in general is quite well investigated. 3988 responding genes were identified. Among them are 22 genes known to be regulated by HS, selected examples are: the heat shock proteins 22A, 22B and 22C, here leading the list of top-regulated genes on place 1 with a FC of 91297 (22A), place 3 with a FC of 45023 (22B) and place 5 with a FC of 6570 (22C). Other examples are several DnaJ-like proteins or the ClpB chaperone, from the Hsp100 family. In total the analysis comprises 8 samples. For heat shock, a 20 ml culture with ~4 M-CM-^W 106 cells per ml in a 100 ml Erlenmeyer flask was incubated at 42M-BM-0C for 45 min in the light with shaking. Control cultures were incubated in the light at 23M-BM-0C (von Gromoff et al., 1989; Kropat et al., 2000). For harvest, cultures were chilled immediately by pouring them onto crushed ice, centrifuged (5 min, 3000 g, 4M-BM-0C) and resuspended in 1 ml TAP. This procedure was repeated three times, resulting in 3 biological replicates.

2011-03-02 | E-GEOD-20859 | biostudies-arrayexpress

Change in gene expression in Chlamydomonas reinhardtii upon heat shock and feeding with hemin and Mg-protoporphyrin

Project description:This SuperSeries is composed of the following subset Series: GSE20859: Change in gene expression in Chlamydomonas reinhardtii upon heat shock GSE20860: Change in gene expression in Chlamydomonas reinhardtii upon feeding with hemin and Mg-protoporphyrin Refer to individual Series

2011-03-11 | E-GEOD-20861 | biostudies-arrayexpress

lincRNAs act in the circuitry controlling pluripotency and differentiation

Project description:While thousands of large intergenic non-coding RNAs (lincRNAs) have been identified in mammals, few have been functionally characterized leading to debate about their biological role. To address this, we performed loss-of-function studies on most lincRNAs expressed in mouse embryonic stem cells (ESC) and characterized the effects on gene expression. Here we show that knockdown of lincRNAs have major consequences on gene expression patterns, comparable to knockdown of well-known ESC regulators. Notably, lincRNAs primarily affect gene expression in trans. We identify dozens of lincRNAs whose knockdown causes an exit from the pluripotent state or upregulation of lineage commitment programs. We integrate lincRNAs into the molecular circuitry of ESCs and show that lincRNA genes are regulated by key transcription factors and that lincRNA transcripts physically bind to multiple chromatin regulatory proteins to affect shared gene expression programs. Together, the results demonstrate that lincRNAs have key roles in the circuitry controlling ESC state. We generated five lentiviral-based shRNAs targeting each of the 237 lincRNAs previously identified in ESCs. These shRNAs successfully targeted 147 lincRNAs and reduced their expression by an average of ~75% compared to endogenous levels in ESCs. As positive controls, we generated shRNAs targeting ~50 genes encoding regulatory proteins, including both transcription factor and chromatin factor genes that have been shown to play critical roles in ESC regulation; we obtained validated hairpins against 40 of these genes. As negative controls, we performed independent infections with lentiviruses containing 27 different shRNAs with no known cellular target RNA.

2011-08-29 | E-GEOD-30245 | biostudies-arrayexpress

Cortical, hippocampal and thalamic whole genome transcriptome data from controls and G72 schizophrenia mice of both sexes

Project description:G72 transgenic (TG, transgenic mice carrying the G72/G30 locus) and wildtype (WT) mice with a CD1 background were used for transcriptome data acquisition. Importantly, three different brain regions, i.e., the retrosplenial cortex, the hippocampus and thalamus, of animals from both sexes were investigated. In total, eight WT control animals (four ♂, age: 23.14 ± 0.00 wks; four ♀, age: 23.46 ± 0.11 wks) and eight G72 transgenic mice (four ♂, age: 23.14 ± 0.00 wks; four ♀, age: 23.14 ± 0.00 wks) were used for dissection of the cortex, thalamus and hippocampus for subsequent transcriptome analysis. Our data allow new insight into relevant alterations in gene transcript levels in G72 mice and allow the reader/user to carry out additional complex analyses to characterize potential sex-, age- and brain-region-specific profiles in gene expression and related pathways. These analyses could trigger biomarker identification and drug research and development in schizophrenia research in the future.

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