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The Z-cad dual fluorescent sensor detects dynamic changes between the epithelial and mesenchymal cellular states.
- Source :
-
BMC biology [BMC Biol] 2016 Jun 17; Vol. 14, pp. 47. Date of Electronic Publication: 2016 Jun 17. - Publication Year :
- 2016
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Abstract
- Background: The epithelial to mesenchymal transition (EMT) has been implicated in metastasis and therapy resistance of carcinomas and can endow cancer cells with cancer stem cell (CSC) properties. The ability to detect cancer cells that are undergoing or have completed EMT has typically relied on the expression of cell surface antigens that correlate with an EMT/CSC phenotype. Alternatively these cells may be permanently marked through Cre-mediated recombination or through immunostaining of fixed cells. The EMT process is dynamic, and these existing methods cannot reveal such changes within live cells. The development of fluorescent sensors that mirror the dynamic EMT state by following the expression of bona fide EMT regulators in live cells would provide a valuable new tool for characterizing EMT. In addition, these sensors will allow direct observation of cellular plasticity with respect to the epithelial/mesenchymal state to enable more effective studies of EMT in cancer and development.<br />Results: We generated a lentiviral-based, dual fluorescent reporter system, designated as the Z-cad dual sensor, comprising destabilized green fluorescent protein containing the ZEB1 3' UTR and red fluorescent protein driven by the E-cadherin (CDH1) promoter. Using this sensor, we robustly detected EMT and mesenchymal to epithelial transition (MET) in breast cancer cells by flow cytometry and fluorescence microscopy. Importantly, we observed dynamic changes in cellular populations undergoing MET. Additionally, we used the Z-cad sensor to identify and isolate minor subpopulations of cells displaying mesenchymal properties within a population comprising predominately epithelial-like cells. The Z-cad dual sensor identified cells with CSC-like properties more effectively than either the ZEB1 3' UTR or E-cadherin sensor alone.<br />Conclusions: The Z-cad dual sensor effectively reports the activities of two factors critical in determining the epithelial/mesenchymal state of carcinoma cells. The ability of this stably integrating dual sensor system to detect dynamic fluctuations between these two states through live cell imaging offers a significant improvement over existing methods and helps facilitate the study of EMT/MET plasticity in response to different stimuli and in cancer pathogenesis. Finally, the versatile Z-cad sensor can be adapted to a variety of in vitro or in vivo systems to elucidate whether EMT/MET contributes to normal and disease phenotypes.
- Subjects :
- Animals
Antigens, CD
Cadherins genetics
Cell Line, Tumor
Green Fluorescent Proteins genetics
Green Fluorescent Proteins metabolism
Humans
Luminescent Proteins genetics
Luminescent Proteins metabolism
Mice
Promoter Regions, Genetic
Transforming Growth Factor beta1 pharmacology
Zinc Finger E-box-Binding Homeobox 1 genetics
Red Fluorescent Protein
Biosensing Techniques
Epithelial Cells cytology
Epithelial-Mesenchymal Transition
Mesenchymal Stem Cells cytology
Subjects
Details
- Language :
- English
- ISSN :
- 1741-7007
- Volume :
- 14
- Database :
- MEDLINE
- Journal :
- BMC biology
- Publication Type :
- Academic Journal
- Accession number :
- 27317311
- Full Text :
- https://doi.org/10.1186/s12915-016-0269-y