Homo sapiens
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA227225 | ENA |
REPOSITORIES: ENA
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Project description:The Hippo pathway is an emerging signaling cascade involved in the regulation of organ size control. It consists of evolutionally conserved protein kinases that are sequentially phosphorylated and activated. The active Hippo pathway subsequently phosphorylates a transcription coactivator, YAP, which precludes its nuclear localization and transcriptional activation. Identification of transcriptional targets of YAP in diverse cellular contexts is therefore critical to the understanding of the molecular mechanisms in which the Hippo pathway restricts tissue growth. We used microarrays to profile the gene expression patterns upon acute siRNA knockdown of Hippo pathway components in multiple mammalian cell lines and identified a set of genes representing immediate transcriptional targets of the Hippo/Yap signaling pathway. Three mammalian cell lines (HEK293T, HepG2, HaCaT) were transfected with scramble siRNA controls or siRNAs against NF2 and LATS2, two core components of the Hippo pathway, simultaneously. Total RNAs were harvested four days after transfection to reveal the gene expression pattern unsing microarry. YAP and TAZ siRNAs were also transfected along with NF2 and LATS2 siRNAs to identify YAP/TAZ-dependent transcriptional targets upon loss of NF2/LATS2.
2013-12-31 | E-GEOD-49384 | biostudies-arrayexpress
Project description:Global microRNA expression analysis in HaCaT human keratinocytes transfected with p72 or control EGFP in addition to siRNAs for NF2 and LATS2.
| PRJNA227229 | ENA
Project description:Anaysis of mRNA changes in HeLa cells following knockdown of Drosha or DGCR8. Drosha is a nuclear RNase III that carries out microRNA (miRNA) processing by cleaving primary microRNA transcript (pri-miRNA). DGCR8 is an essential co-factor of Drosha. Experiment Overall Design: siRNA against Drosha or DGCR8 were trasnfected into HeLa cells. siRNA against GFP was used as a control. Biologically duplicated total RNAs were prepared from HeLa cells, 24 hrs and 48 hrs after siRNA transfection.
2009-02-21 | E-GEOD-13639 | biostudies-arrayexpress
Project description:This transcriptome study is intended to discover the effects of MYC and TP63 on human keratinocytes differentiation at the genome level. Human keratinocyte cell line, HaCaT cells were transfected by siRNAs with the help of transfection reagent, INTERFERin. 48 hours post-transfection, total RNA was extracted from cells. We used a non-targeting siRNA as a negative control. siRNA targeting MYC or TP63 was obtained from QIAGEN, and RNAs from wild type HaCaT cells were used as reference samples. We did 3 biological replicates and 2 'dye swap' for each sample. Totally we got 12 samples, with 2 dye-swap for each sample, and 48 arrays used for all hybridization.
2010-05-15 | E-GEOD-17394 | biostudies-arrayexpress
Project description:Global downregulation of microRNAs (miRNAs) is commonly observed in human cancers and can have a causative role in tumorigenesis. The mechanisms responsible for this phenomenon remain poorly understood. Here we show that YAP, the downstream target of the tumor-suppressive Hippo signaling pathway regulates miRNA biogenesis in a cell density-dependent manner. At low cell density, nuclear YAP binds and sequesters p72 (DDX17), a regulatory component of the miRNA processing machinery. At high cell density, Hippo-mediated cytoplasmic retention of YAP facilitates p72 association with Microprocessor and binding to a specific sequence motif in pri-miRNAs. Inactivation of the Hippo pathway or expression of constitutively active YAP causes widespread miRNA suppression in cells and tumors and a corresponding post-transcriptional induction of MYC expression. Thus, the Hippo pathway links contact-inhibition regulation to miRNA biogenesis and may be responsible for the widespread miRNA repression observed in cancer. Two conditions (transfection of p72 or control EGFP in addition to siRNAs for NF2 and LATS2) were analyzed in duplicate.
2014-03-03 | E-GEOD-52205 | biostudies-arrayexpress
Project description:Global microRNA expression analysis in HaCaT human keratinocytes transfected with siRNA for YAP or negative control
| PRJNA227226 | ENA
Project description:Global downregulation of microRNAs (miRNAs) is commonly observed in human cancers and can have a causative role in tumorigenesis. The mechanisms responsible for this phenomenon remain poorly understood. Here we show that YAP, the downstream target of the tumor-suppressive Hippo signaling pathway regulates miRNA biogenesis in a cell density-dependent manner. At low cell density, nuclear YAP binds and sequesters p72 (DDX17), a regulatory component of the miRNA processing machinery. At high cell density, Hippo-mediated cytoplasmic retention of YAP facilitates p72 association with Microprocessor and binding to a specific sequence motif in pri-miRNAs. Inactivation of the Hippo pathway or expression of constitutively active YAP causes widespread miRNA suppression in cells and tumors and a corresponding post-transcriptional induction of MYC expression. Thus, the Hippo pathway links contact-inhibition regulation to miRNA biogenesis and may be responsible for the widespread miRNA repression observed in cancer. Four conditions (siCtrl, si p72, siNF2/LATS2 and siDROSHA/DGCR8) were analyzed in duplicate.
2014-03-03 | E-GEOD-52186 | biostudies-arrayexpress
Project description:Drosha is a type III RNAse, which plays a critical role in miRNA biogenesis. Drosha and its double-stranded RNA-binding partner protein Pasha/DGCR8 likely recognize and cleave miRNA precursor RNAs or pri-miRNA hairpins co-transcriptionally. To identify RNAs processed by Drosha, we used tiling microarrays to examine transcripts after depletion of drosha mRNA with dsRNA in Drosophila Schneider S2 cells. This strategy identified 137 Drosha-regulated RNAs, including 11 putative pri-miRNAs comprising 15 annotated miRNAs. Most of the identified pri-miRNAs seem extremely large, >10 kilobases as revealed by both the Drosha knock down strategy and by RNA PolII chromatin IP followed by Drosophila tiling microarrays. Surprisingly, more than a hundred additional RNAs not annotated as miRNAs are under Drosha control and are likely to be direct targets of Drosha action. This is because many of them encode annotated genes, and unlike bona fide pri-miRNAs, they are not affected by depletion of the miRNA processing factor, dicer-1. Moreover, application of the evofold analysis software indicates that at least 25 of the Drosha-regulated RNAs contain evolutionarily conserved hairpins similar to those recognized by the Drosha-Pasha/DGCR8 complex in pri-miRNAs. One of these hairpins is located in the 5′ UTR of both pasha and mammalian DGCR8. These observations suggest that a negative feedback loop acting on pasha mRNA may regulate the miRNA-biogenesis pathway: i.e., excess Drosha cleaves pasha/DGCR8 primary transcripts and leads to a reduction in pasha/DGCR8 mRNA levels and Pasha/DGCR8 synthesis. Keywords: time course, ChIP-chip
2008-12-25 | GSE14215 | GEO
Project description:To examine the effect of DEAD-box RNA helicase subunits on the primary miRNAs processing by Drosha complex, we made knockout mice of p72, DEAD-box RNA helicase, a component of Drosha complex. And we compare the miRNA expression profiles derived from whole mice embryo between wild-type and p72 KO mice.
2007-04-12 | E-MEXP-1052 | biostudies-arrayexpress
Project description:Regulation of microRNA (miR) biogenesis is complex and stringently controlled. Here we identify the kinase GSK3B as an important modulator of global miR biogenesis at microprocessor level. Repression of GSK3B activity reduces Drosha activity towards pri-miRs, leading to accumulation of unprocessed pri-miRs and reduction of pre-miRs and mature miRs without altering levels or cellular localisationof miR biogenesisproteins. Conversly, GSK3B activation increases Drosha activityand mature miR accumulation. GSK3B achieves this through promoting Drosha:cofactor and Drosha:pri-miR interactions: it binds to DGCR8 and p72 in the Microprocessor, an effect dependent of RNA. Indeed GSK3B itself can immunoprecipitate pri-miRs, suggesting possible RNA binding capacity. Kinase assays identify the mechanism for GSK3B-enhansed Drosha activity, which requires GSK3B nuclear localisation, as phosphorylation of Drosha at s300 and/or s302 confirmed by enhanced Drosha activity and association with cofactors, and increased abundance of mature microRNAs in the presence of phospho-mimetic Drosha. Functional implicatons of GSK3B-enhanced miR biogenesis are illustrated by increased levels of GSK3B-upregulatd miR targets following GSK3B inhibition. These data, the first link GSK3B with the miR cascade in humans, hilight a novel pro-biogenesis role for GSK3B in increasing miR biogenesis as a component of the Microprocessor complex with wide ranging consiquences.
2016-05-20 | GSE81141 | GEO