PUBLICATION
High-sensitivity measurements of multiple kinase activities in live single cells
- Authors
- Regot, S., Hughey, J.J., Bajar, B.T., Carrasco, S., Covert, M.W.
- ID
- ZDB-PUB-230526-71
- Date
- 2014
- Source
- Cell 157: 172417341724-34 (Journal)
- Registered Authors
- Keywords
- none
- MeSH Terms
-
- Sequence Alignment
- JNK Mitogen-Activated Protein Kinases/chemistry
- JNK Mitogen-Activated Protein Kinases/metabolism
- Mice
- Molecular Sequence Data
- Single-Cell Analysis
- Phosphotransferases/metabolism*
- Animals
- Amino Acid Sequence
- Biosensing Techniques/methods*
- PubMed
- 24949979 Full text @ Cell
Citation
Regot, S., Hughey, J.J., Bajar, B.T., Carrasco, S., Covert, M.W. (2014) High-sensitivity measurements of multiple kinase activities in live single cells. Cell. 157:172417341724-34.
Abstract
Increasing evidence has shown that population dynamics are qualitatively different from single-cell behaviors. Reporters to probe dynamic, single-cell behaviors are desirable yet relatively scarce. Here, we describe an easy-to-implement and generalizable technology to generate reporters of kinase activity for individual cells. Our technology converts phosphorylation into a nucleocytoplasmic shuttling event that can be measured by epifluorescence microscopy. Our reporters reproduce kinase activity for multiple types of kinases and allow for calculation of active kinase concentrations via a mathematical model. Using this technology, we made several experimental observations that had previously been technicallyunfeasible, including stimulus-dependent patterns of c-Jun N-terminal kinase (JNK) and nuclear factor kappa B (NF-κB) activation. We also measured JNK, p38, and ERK activities simultaneously, finding that p38 regulates the peak number, but not the intensity, of ERK fluctuations. Our approach opens the possibility of analyzing a wide range of kinase-mediated processes in individual cells.
Genes / Markers
Expression
Phenotype
Mutations / Transgenics
Human Disease / Model
Sequence Targeting Reagents
Fish
Orthology
Engineered Foreign Genes
Mapping