A fluorescent reporter of AMPK activity and cellular energy stress

Cell Metab. 2011 Apr 6;13(4):476-486. doi: 10.1016/j.cmet.2011.03.006.

Abstract

AMP-activated protein kinase (AMPK) is activated when the AMP/ATP ratio in cells is elevated due to energy stress. Here, we describe a biosensor, AMPKAR, that exhibits enhanced fluorescence resonance energy transfer (FRET) in response to phosphorylation by AMPK, allowing spatiotemporal monitoring of AMPK activity in single cells. We show that this reporter responds to a variety of stimuli that are known to induce energy stress and that the response is dependent on AMPK α1 and α2 and on the upstream kinase LKB1. Interestingly, we found that AMPK activation is confined to the cytosol in response to energy stress but can be observed in both the cytosol and nucleus in response to calcium elevation. Finally, using this probe with U2OS cells in a microfluidic device, we observed a very high cell-to-cell variability in the amplitude and time course of AMPK activation and recovery in response to pulses of glucose deprivation.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • AMP-Activated Protein Kinases / metabolism*
  • Amino Acid Sequence
  • Calcium / metabolism
  • Calcium-Calmodulin-Dependent Protein Kinase Kinase / metabolism
  • Cell Line, Tumor
  • Energy Metabolism
  • Fluorescence Resonance Energy Transfer
  • Humans
  • Microfluidic Analytical Techniques
  • Molecular Sequence Data
  • Phosphorylation
  • Protein-Serine-Threonine Kinases / metabolism

Substances

  • STK11 protein, human
  • PRKAA1 protein, human
  • PRKAA2 protein, human
  • Protein-Serine-Threonine Kinases
  • Calcium-Calmodulin-Dependent Protein Kinase Kinase
  • AMP-Activated Protein Kinases
  • Calcium