Cellular Application of Genetically Encoded Sensors and Impeders of AMPK.
Unraveling the spatiotemporal dynamics of 5'-AMP-activated protein kinase (AMPK) signaling is necessary to bridge the gap between nutrient signaling and downstream function. Three genetically encoded Förster Resonance Energy Transfer (FRET)-based AMPK biosensors are available yielding insight into how AMPK-derived signal propagates throughout a cell in response to particular inputs. These findings, together with accumulating evidence obtained from biochemical techniques, promise to give a holistic understanding of the AMPK signaling. In this protocol, we describe the procedures and materials required for imaging intracellular AMPK activity in an organelle-specific manner, with a focus on ABKAR, a FRET-based biosensor. In addition, we introduce a novel AMPK inhibitor peptide that allows us to inhibit AMPK activity at specific subcellular compartments.
['Miyamoto|Takafumi|T|', 'Rho|Elmer|E|', 'Kim|Allen|A|', 'Inoue|Takanari|T|']
D055372:AMP-Activated Protein Kinases D000818:Animals D015374:Biosensing Techniques D019556:COS Cells D002522:Chlorocebus aethiops D005347:Fibroblasts D031541:Fluorescence Resonance Energy Transfer D006367:HeLa Cells D006801:Humans D000069416:Intravital Microscopy D008164:Luminescent Proteins D051379:Mice D008856:Microscopy, Fluorescence D057054:Molecular Imaging D015388:Organelles D015398:Signal Transduction D012984:Software
Biosensor Organelle-specific Inhibitor peptide Compartmentalization Förster resonance energy transfer (FRET) Kinase activity monitoring Signaling dynamics