TY - JOUR
T1 - A genetically encoded fluorescent sensor of ERK activity
AU - Harvey, Christopher D.
AU - Ehrhardt, Anka G.
AU - Cellurale, Cristina
AU - Zhong, Haining
AU - Yasuda, Ryohei
AU - Davis, Roger J.
AU - Svoboda, Karel
PY - 2008/12/9
Y1 - 2008/12/9
N2 - 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.
AB - 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.
KW - FRET
KW - Fluorescence lifetime imaging microscopy
KW - MAPK
UR - http://www.scopus.com/inward/record.url?scp=58049221116&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=58049221116&partnerID=8YFLogxK
U2 - 10.1073/pnas.0804598105
DO - 10.1073/pnas.0804598105
M3 - Article
C2 - 19033456
AN - SCOPUS:58049221116
SN - 0027-8424
VL - 105
SP - 19264
EP - 19269
JO - Proceedings of the National Academy of Sciences of the United States of America
JF - Proceedings of the National Academy of Sciences of the United States of America
IS - 49
ER -