High-sensitivity measurements of multiple kinase activities in live single cells

Sergio Regot Rodriguez De Mier, Jacob J. Hughey, Bryce T. Bajar, Silvia Carrasco, Markus W. Covert

Research output: Contribution to journalArticle

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.

Original languageEnglish (US)
Pages (from-to)1724-1734
Number of pages11
JournalCell
Volume157
Issue number7
DOIs
StatePublished - Jun 19 2014
Externally publishedYes

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Phosphotransferases
JNK Mitogen-Activated Protein Kinases
Technology
Population dynamics
Phosphorylation
NF-kappa B
Population Dynamics
Microscopy
Microscopic examination
Theoretical Models
Chemical activation
Mathematical models

ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology(all)

Cite this

High-sensitivity measurements of multiple kinase activities in live single cells. / Regot Rodriguez De Mier, Sergio; Hughey, Jacob J.; Bajar, Bryce T.; Carrasco, Silvia; Covert, Markus W.

In: Cell, Vol. 157, No. 7, 19.06.2014, p. 1724-1734.

Research output: Contribution to journalArticle

Regot Rodriguez De Mier, Sergio ; Hughey, Jacob J. ; Bajar, Bryce T. ; Carrasco, Silvia ; Covert, Markus W. / High-sensitivity measurements of multiple kinase activities in live single cells. In: Cell. 2014 ; Vol. 157, No. 7. pp. 1724-1734.
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