UMMS Affiliation

Program in Molecular Medicine

Publication Date

2008-12-09

Document Type

Article

Disciplines

Biochemistry | Biophysics | Cell Biology | Cellular and Molecular Physiology | Molecular Biology

Abstract

The activity of the ERK has complex spatial and temporal dynamics that are important for the specificity of downstream effects. However, current biochemical techniques do not allow for the measurement of ERK signaling with fine spatiotemporal resolution. We developed a genetically encoded, FRET-based sensor of ERK activity (the extracellular signal-regulated kinase activity reporter, EKAR), optimized for signal-to-noise ratio and fluorescence lifetime imaging. EKAR selectively and reversibly reported ERK activation in HEK293 cells after epidermal growth factor stimulation. EKAR signals were correlated with ERK phosphorylation, required ERK activity, and did not report the activities of JNK or p38. EKAR reported ERK activation in the dendrites and nucleus of hippocampal pyramidal neurons in brain slices after theta-burst stimuli or trains of back-propagating action potentials. EKAR therefore permits the measurement of spatiotemporal ERK signaling dynamics in living cells, including in neuronal compartments in intact tissues.

Keywords

fluorescence lifetime imaging microscopy, FRET, MAPK

Rights and Permissions

Publisher PDF posted as allowed by the publisher's author rights policy at http://www.pnas.org/site/aboutpnas/authorfaq.xhtml.

DOI of Published Version

10.1073/pnas.0804598105

Source

Proc Natl Acad Sci U S A. 2008 Dec 9;105(49):19264-9. doi: 10.1073/pnas.0804598105. Epub 2008 Nov 25. Link to article on publisher's site

Journal/Book/Conference Title

Proceedings of the National Academy of Sciences of the United States of America

Related Resources

Link to Article in PubMed

PubMed ID

19033456

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