An ultrasensitive biosensor for high-resolution kinase activity imaging in awake mice.
Alprostadil
/ pharmacology
Animals
Biosensing Techniques
Cyclic AMP-Dependent Protein Kinases
/ genetics
Dihydroxyphenylalanine
/ pharmacology
Dinoprostone
/ pharmacology
Fluorescent Dyes
/ chemistry
Gene Expression
Gene Library
Genes, Reporter
Glucagon-Like Peptide 1
/ pharmacology
HEK293 Cells
HeLa Cells
High-Throughput Screening Assays
Hippocampus
/ cytology
Humans
Mice
Microscopy, Fluorescence, Multiphoton
Myocytes, Cardiac
/ drug effects
Neurons
/ drug effects
Optical Imaging
/ methods
Primary Cell Culture
Signal Transduction
Journal
Nature chemical biology
ISSN: 1552-4469
Titre abrégé: Nat Chem Biol
Pays: United States
ID NLM: 101231976
Informations de publication
Date de publication:
01 2021
01 2021
Historique:
received:
28
01
2020
accepted:
21
08
2020
pubmed:
30
9
2020
medline:
20
2
2021
entrez:
29
9
2020
Statut:
ppublish
Résumé
Protein kinases control nearly every facet of cellular function. These key signaling nodes integrate diverse pathway inputs to regulate complex physiological processes, and aberrant kinase signaling is linked to numerous pathologies. While fluorescent protein-based biosensors have revolutionized the study of kinase signaling by allowing direct, spatiotemporally precise kinase activity measurements in living cells, powerful new molecular tools capable of robustly tracking kinase activity dynamics across diverse experimental contexts are needed to fully dissect the role of kinase signaling in physiology and disease. Here, we report the development of an ultrasensitive, second-generation excitation-ratiometric protein kinase A (PKA) activity reporter (ExRai-AKAR2), obtained via high-throughput linker library screening, that enables sensitive and rapid monitoring of live-cell PKA activity across multiple fluorescence detection modalities, including plate reading, cell sorting and one- or two-photon imaging. Notably, in vivo visual cortex imaging in awake mice reveals highly dynamic neuronal PKA activity rapidly recruited by forced locomotion.
Identifiants
pubmed: 32989297
doi: 10.1038/s41589-020-00660-y
pii: 10.1038/s41589-020-00660-y
pmc: PMC7773213
mid: NIHMS1654070
doi:
Substances chimiques
Fluorescent Dyes
0
Dihydroxyphenylalanine
63-84-3
Glucagon-Like Peptide 1
89750-14-1
Cyclic AMP-Dependent Protein Kinases
EC 2.7.11.11
Alprostadil
F5TD010360
Dinoprostone
K7Q1JQR04M
Types de publication
Journal Article
Research Support, N.I.H., Extramural
Research Support, U.S. Gov't, Non-P.H.S.
Langues
eng
Sous-ensembles de citation
IM
Pagination
39-46Subventions
Organisme : NIDDK NIH HHS
ID : R01 DK073368
Pays : United States
Organisme : NINDS NIH HHS
ID : F31 NS108593
Pays : United States
Organisme : NINDS NIH HHS
ID : U24 NS109107
Pays : United States
Organisme : NIMH NIH HHS
ID : R01 MH111516
Pays : United States
Organisme : NIMH NIH HHS
ID : DP2 MH107056
Pays : United States
Organisme : NINDS NIH HHS
ID : U01 NS094246
Pays : United States
Organisme : NCI NIH HHS
ID : R35 CA197622
Pays : United States
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