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Differential expression of GFP+ and RFP+ TriPyMT cells

ABSTRACT: Tri-PyMT cell is a breast tumor cell line established from Tri-PyMT (MMTV-PyMT/fsp1-Cre/Rosa26-RGFP) primary breast tumors. The cells switch from RFP+ to GFP+ during epithelial to mesenchymal transition (EMT). Differential expressed genes between EMT and non-EMT tumor cells will reveal interesting targets for anti-EMT approaches.

ORGANISM(S): Mus musculus

PROVIDER: GSE72480 | GEO | 2015/09/01

SECONDARY ACCESSION(S): PRJNA294121

REPOSITORIES: GEO

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Differential expression of GFP+ and RFP+ TriPyMT cells

Project description:Tri-PyMT cell is a breast tumor cell line established from Tri-PyMT (MMTV-PyMT/fsp1-Cre/Rosa26-RGFP) primary breast tumors. The cells switch from RFP+ to GFP+ during epithelial to mesenchymal transition (EMT). Differential expressed genes between EMT and non-EMT tumor cells will reveal interesting targets for anti-EMT approaches. Tri-PyMT cells were culture in medium with serum (10%FBS). Cells were treated with or without 4-hydroperoxy-cyclophosphamide (4µM) for 7 days and sorted by flow cytometry into RFP+ and GFP+ subpopulations for RNA extraction and subsequent RNA-Seq analysis.

2015-09-01 | E-GEOD-72480 | biostudies-arrayexpress

Single cell sequencing of Tri-PyMT cells

Project description:To study the contribution of epithelial-to-mesenchymal transition (EMT) in tumor metastasis, we established an EMT lineage tracing model in a multiple-transgenic mouse (MMTV-PyMT/Fsp1-Cre/Rosa26mT/mG, Tri-PyMT). In this model, breast tumor cells that underwent EMT would irreversibly switch their expression of the fluorescent marker from RFP+ to GFP+ due to the mesenchymal specific Cre expression. Surprisingly, we found that metastatic tumor cells did not convert from RFP+ to GFP+. Lung metastases were predominantly composed of RFP+ tumor cells persistently exhibiting epithelial phenotypes. These findings challenge the concept that EMT is required for metastasis, and have attracted vigorous discussion about its true contributions to metastasis. One of the major concerns with the EMT lineage tracing model is that expression of GFP only indicates the complete EMT transition. The partial-EMT programming, a novel concept describing tumor cells exhibiting both epithelial and mesenchymal features, could possibly fail to launch the Fsp1-Cre-mediated fluorescent marker switch. To address this concern, we evaluated the EMT status of Tri-PyMT cells on the single cell level by performing single cell RNA-sequencing (scRNA-seq). We found that the expression of epithelial markers (i.e. Epcam and Krt18) and mesenchymal markers (i.e. Vim and S110a4) were largely confined to RFP+ and GFP+ subpopulations, respectively. Based on the differentially expressed EMT related genes, we observed an EMT spectrum in which the low EMT score end was enriched with RFP+ cells, whereas the high EMT score end was enriched with GFP+ cells. Indeed, the fluorescent marker switch in Tri-PyMT cells predicted their EMT status with high specificity and sensitivity. Tri-PyMT cells which were derived from the primary breast tumor of triple transgenic mouse (MMTV-PyMT:Fsp1-cre:Rosa26mT/mG) were sorted into RFP+ or GFP+ populations by FACS and subjected for Drop-seq analysis.

2022-02-01 | GSE126374 | GEO

Sorted MDMs with RFP+GFP+ or RFP+GFP- Mtb

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Sorted MDMs with RFP+GFP+ or RFP+GFP- Mtb

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HEK293 GFP-LacZ-E2F-RFP

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Differential Gene expression in the mouse mammary tumors of PyMT-Malat1 wild-type (WT), PyMT-Malat1 knockout (KO) and PyMT-Malat1 knockout with Malat1 transgene expression (TG)

Project description:Previously, lncRNA Malat1 knockout mice were generated by insertional inactivation. By crossing this line to MMTV-PyMT mammary tumor mouse model, we produced PyMT;Malat1 wild-type (WT) and PyMT;Malat1 knockout (KO). Furthermore, we generated Malat1 transgenic mice by targeting ROSA26 locus and bred them to PyMT;Malat1 knockout mice to produce Malat1-rescued PyMT;Malat1 knockout;Malat1 transgenic animals (TG). Using mammary tumors from the three groups of animals, we performed RNA-Seq analysis to identify differentially up-regulated genes in KO tumors to find novel target genes of YAP-TEAD pathway.

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PyMT-1099, a murine derived cell line is a breast cancer specific model for EMT

Project description:Background: Epithelial-mesenchymal transition (EMT) has been implicated in metastasis, drug resistance, survival under stress and also conferring stem cell-like traits to cancer cells. However, several of the studies have been carried out using model systems that don’t appropriately recapitulate all stages of the dynamic process of EMT. Hence, there is a need to overcome this limitation by development of a model system that allows us to mimic each stage of EMT and accurately assess the plastic changes associated with it. Methods: We have derived a cancer cell line from the PyMT-MMTV model of breast cancer, named PyMT-1099 cells, and undertaken a detailed characterization of the morpho-genetic changes it undergoes during a TGF-induced EMT. Further, we have also performed high throughput transcriptomics on PyMT-1099 cells undergoing EMT in a high resolution kinetic of TGF treatment. Results: We show that PyMT-1099 cells undergo an EMT comparable to the classically used immortalized NMuMG cells as assessed by morphological, and marker expression changes on TGFtreatment. Further, PyMT-1099 cells can also migrate in vitro in response to TGF treatment. These cells are also tumorigenic and lead to metastasis formation when transplanted into immunocompromised mice. Conclusion: In this study we report the development of PyMT-1099 cells as an excellent tool to model and study breast cancer-associated EMT both in vitro and in vivo and show that these cells overcome the limitations posed by other cellular systems currently being used to study EMT

2018-10-03 | GSE114572 | GEO

Mesenchymal Epithelial Transition (MET) Timecourse of NMuMG-E9 cells treated with TGFb

Project description:Background: Epithelial-mesenchymal transition (EMT) has been implicated in metastasis, drug resistance, survival under stress and also conferring stem cell-like traits to cancer cells. However, several of the studies have been carried out using model systems that don’t appropriately recapitulate all stages of the dynamic process of EMT. Hence, there is a need to overcome this limitation by development of a model system that allows us to mimic each stage of EMT and accurately assess the plastic changes associated with it. Methods: We have derived a cancer cell line from the PyMT-MMTV model of breast cancer, named PyMT-1099 cells, and undertaken a detailed characterization of the morpho-genetic changes it undergoes during a TGFbeta-induced EMT. Further, we have also performed high throughput transcriptomics on PyMT-1099 cells undergoing EMT in a high resolution kinetic of TGFbeta treatment. Results: We show that PyMT-1099 cells undergo an EMT comparable to the classically used immortalized NMuMG cells as assessed by morphological, and marker expression changes on TGFbeta treatment. Further, PyMT-1099 cells can also migrate in vitro in response to TGFbeta treatment. These cells are also tumorigenic and lead to metastasis formation when transplanted into immunocompromised mice. Conclusion: In this study we report the development of PyMT-1099 cells as an excellent tool to model and study breast cancer-associated EMT both in vitro and in vivo and show that these cells overcome the limitations posed by other cellular systems currently being used to study EMT

2018-10-03 | GSE117474 | GEO

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