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Dynamic Chromatin Modification Sustains Epithelial-Mesenchymal Transition following Inducible Expression of Snail-1 (ChIP-chip)

ABSTRACT: Epithelial-Mesenchymal Transition (EMT) is thought to contribute to cancer metastasis, but its underlying mechanisms are not well understood. To define early steps in this cellular transformation, we analyzed human mammary epithelial cells with tightly regulated expression of Snail-1, a master regulator of EMT. Following Snail-1 induction, epithelial markers were repressed within 6 hours and mesenchymal genes induced at 24 hours. Snail-1 binding to its target promoters was transient (6-48 hours) despite continued protein expression and it was followed by both transient and long-lasting chromatin changes. To generate a potent reversible EMT-inducing stimulus, we created a Snail-1 retroviral expression construct, using a fused estrogen receptor (ER) response element to mediate regulation by exogenous 4-hydroxy-tamoxifen (4-OHT). Since Snail-1 protein stability and nuclear localization are suppressed by GSK3-beta-mediated phosphorylation, we substituted the six targeted amino acids (ER-Snail-1(6SA)), thus conferring constitutive activity to the induced protein (Zhou et al., 2004, Pubmed ID 15448698). Infection of non-transformed, immortalized human mammary epithelial MCF10A cells with ER-Snail-1(6SA), followed by treatment with 4-OHT, triggered morphological and biomarker characteristics of EMT. At 0, 6, 48 and 120 hours after beginning exposure to 4-OHT in ethanol (or, for controls, ethanol only) we ChIPed Snail-1 and 6 histone marks. We perfomed two replicates of each, except we only had one replicate of H3K27Me3 at 6 hours.

ORGANISM(S): Homo sapiens

SUBMITTER: Ben Wittner

PROVIDER: E-GEOD-52591 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Dynamic Chromatin Modification Sustains Epithelial-Mesenchymal Transition following Inducible Expression of Snail-1 (gene expression)

Project description:Epithelial-Mesenchymal Transition (EMT) is thought to contribute to cancer metastasis, but its underlying mechanisms are not well understood. To define early steps in this cellular transformation, we analyzed human mammary epithelial cells with tightly regulated expression of Snail-1, a master regulator of EMT. Following Snail-1 induction, epithelial markers were repressed within 6 hours and mesenchymal genes induced at 24 hours. Snail-1 binding to its target promoters was transient (6-48 hours) despite continued protein expression and it was followed by both transient and long-lasting chromatin changes. To generate a potent reversible EMT-inducing stimulus, we created a Snail-1 retroviral expression construct, using a fused estrogen receptor (ER) response element to mediate regulation by exogenous 4-hydroxy-tamoxifen (4-OHT). Since Snail-1 protein stability and nuclear localization are suppressed by GSK3-beta-mediated phosphorylation, we substituted the six targeted amino acids (ER-Snail-1(6SA)), thus conferring constitutive activity to the induced protein (Zhou et al., 2004, Pubmed ID 15448698). Infection of non-transformed, immortalized human mammary epithelial MCF10A cells with ER-Snail-1(6SA), followed by treatment with 4-OHT, triggered morphological and biomarker characteristics of EMT. At 3, 6, 12, 24, 72 and 120 hours after beginning exposure to 4-OHT in ethanol (or, for controls, ethanol only) we extracted RNA and did gene expression analysis using microarrays. We perfomed three replicates of each.

2013-12-18 | E-GEOD-52592 | biostudies-arrayexpress

Dynamic Chromatin Modification Sustains Epithelial-Mesenchymal Transition following Inducible Expression of Snail-1

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

2013-12-18 | E-GEOD-52593 | biostudies-arrayexpress

Expression data of mammary epithelial cells during the epithelial-mesenchymal transition

Project description:The EMT program allows epithelial cells to become endowed with motility, invasiveness and stem cell traits. We investigated difference in signaling networks that are differentially utilized in EMTed and non-EMTed cells, thereby identifying therapeutic targets that are unique to EMT/cancer stem cells. We expressed the EMT transcription factors, Twist, Snail and Slug in HMLE human mammary epithelial cells, and compared their gene expression with parental cells. We identified kinases that are more differentially regulated between the epithelial and mesenchymal cell state.

2013-09-09 | E-GEOD-43495 | biostudies-arrayexpress

Retinoic acid induces Snai1 in stem cells of the preimplantation blastocyst to initiate differentiation

Project description:Although much is known about the pluripotency self-renewal circuitry, the molecular events that lead embryonic stem cells (ESCs) exit from pluripotency and begin differentiation are largely unknown. We found that the zinc finger transcription factor Snai1, involved in gastrulation and epithelial- mesenchymal transition (EMT) is already expressed in the inner cell mass of the preimplantation blastocysts. In ESCs Snai1 does not respond to TGFÎ± or BMP4 signalling but it is induced by retinoic acid (RA) treatment, which induces the binding, on the Snai1 promoter, of the retinoid receptors RARÎ³ and RXRÎ± the dissociation of the Polycomb repressor compex 2 (PRC2) which results in the decrease of H3K27me3 and the increase of histone H3K4me3. Snai1 mediates the repression of pluripotency genes by binding directly to the promoters of Nanog, Nr5a2, Tcl1, c-Kit, and Tcfcp2l1. The transient activation of Snai1 in embryoid bodies induces the expression of the markers of all three germ layers. These results suggest that Snai1 is a key factor that triggers ESCs exit from the pluripotency state and initiate their differentiation processes. microarray analysis of embryonic stem cells (ESC) expressing Snail-ER at various time points of induction with 4-OHT

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The mRNA expression of FaDu-Ctrl, FaDu-Snail, FaDu-Snail2R cells

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2014-08-16 | E-GEOD-58592 | biostudies-arrayexpress

Circulating Breast Tumor Cells Exhibit Dynamic Changes in Epithelial and Mesenchymal Cell Composition

Project description:By applying RNA-ISH and RNAseq to circulating tumor cells (CTCs), the study provides definitive evidence of epithelial to mesenchymal transition (EMT) across all histological types of breast cancer, identifying mediators such as FOXC1 and TGF-β signaling, and demonstrating dynamic treatment-associated changes in EMT within clusters of CTCs. Epithelial to mesenchymal transition (EMT) has been postulated to contribute to the migration and dissemination of cancer cells, but supporting histopathological evidence is limited. We used a microfluidic device to isolate circulating tumor cells (CTCs), combined with multiplex fluorescent RNA-in-situ hybridization (ISH) and RNA sequencing, to quantify and characterize EMT in breast cancer cells within the bloodstream. Whereas only rare (0.1-10%) cells in the primary tumor expressed both mesenchymal and epithelial markers, such biphenotypic as well as purely mesenchymal cells were enriched among CTCs, across all histological subtypes of breast cancer. In an index patient followed longitudinally, fluctuation in epithelial and mesenchymal states was observed as a function of initial response and subsequent resistance to therapy. Mesenchymal markers were predominant in clusters of tumor cells, many of which had adherent platelets. Finally, RNA sequencing of CTC clusters identified TGF-β and other EMT-related signatures, which were absent from more epithelial CTCs. FOXC1, a known regulator of EMT, was abundantly expressed in mesenchymal CTCs and was detectable within localized regions of the primary breast tumor. Together, these data support a role for EMT in the blood-borne dissemination of breast cancer and point to the dynamic nature of this cell fate change.

2013-02-01 | E-GEOD-41245 | biostudies-arrayexpress

Dynamic Chromatin Modification Sustains Epithelial-Mesenchymal Transition following Inducible Expression of Snail-1 (ChIP-chip)

2013-12-18 | GSE52591 | GEO

Dynamic Chromatin Modification Sustains Epithelial-Mesenchymal Transition following Inducible Expression of Snail-1 (gene expression)

2013-12-18 | GSE52592 | GEO

Roles of Snail-regulated miRNAs in colon cancer progression

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2013-11-20 | E-GEOD-39286 | biostudies-arrayexpress

Network Understanding of SNAIL inhibitor induced perturbations in EMT models

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