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Live visualization and quantification of pathway signaling with dual fluorescent and bioluminescent reporters
- Source :
- Biochemical and Biophysical Research Communications. 448:281-286
- Publication Year :
- 2014
- Publisher :
- Elsevier BV, 2014.
-
Abstract
- Despite their fundamental importance, the dynamics of signaling pathways in living cells remain challenging to study, due to a lack of non-invasive tools for temporal assessment of signal transduction in desired cell models. Here we report a dual-reporter strategy that enables researchers to monitor signal transduction in mammalian cells in real-time, both temporally and quantitatively. This is achieved by co-expressing green fluorescent protein and firefly luciferase in response to signaling stimuli. To display the versatility of this approach, we constructed and assessed eight unique signaling pathway reporters. We further validated the system by establishing stable NF-κB pathway reporter cell lines. Using these stable cell lines, we monitored the activity of NF-κB-mediated inflammatory pathway in real-time, both visually and quantitatively. Live visualization has the power to reveal individual cell responses and is compatible with single cell analysis, In addition, we provide evidence that this system is readily amenable to a high-throughput format. Together, our findings demonstrate the potential of the dual reporter system, which significantly improves the capacity to study signal transduction pathways in mammalian cells.
- Subjects :
- Green Fluorescent Proteins
Cell
Biophysics
Biology
Response Elements
Transfection
Biochemistry
Green fluorescent protein
Single-cell analysis
Genes, Reporter
Luciferases, Firefly
medicine
Humans
Bioluminescence
Luciferase
Molecular Biology
Transcription factor
NF-kappa B
Cell Biology
Recombinant Proteins
High-Throughput Screening Assays
Cell biology
HEK293 Cells
medicine.anatomical_structure
Cell culture
Single-Cell Analysis
Signal transduction
Signal Transduction
Subjects
Details
- ISSN :
- 0006291X
- Volume :
- 448
- Database :
- OpenAIRE
- Journal :
- Biochemical and Biophysical Research Communications
- Accession number :
- edsair.doi.dedup.....2b313e4a9ea5bc81128728db9ce8f98e