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

Sergi Regot, 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

ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology(all)

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    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(7), 1724-1734. https://doi.org/10.1016/j.cell.2014.04.039