Fluorescence-based ATG8 sensors monitor localization and function of LC3/GABARAP proteins

EMBO J. 2017 Feb 15;36(4):549-564. doi: 10.15252/embj.201695063. Epub 2016 Dec 27.


Autophagy is a cellular surveillance pathway that balances metabolic and energy resources and transports specific cargos, including damaged mitochondria, other broken organelles, or pathogens for degradation to the lysosome. Central components of autophagosomal biogenesis are six members of the LC3 and GABARAP family of ubiquitin-like proteins (mATG8s). We used phage display to isolate peptides that possess bona fide LIR (LC3-interacting region) properties and are selective for individual mATG8 isoforms. Sensitivity of the developed sensors was optimized by multiplication, charge distribution, and fusion with a membrane recruitment (FYVE) or an oligomerization (PB1) domain. We demonstrate the use of the engineered peptides as intracellular sensors that recognize specifically GABARAP, GABL1, GABL2, and LC3C, as well as a bispecific sensor for LC3A and LC3B. By using an LC3C-specific sensor, we were able to monitor recruitment of endogenous LC3C to Salmonella during xenophagy, as well as to mitochondria during mitophagy. The sensors are general tools to monitor the fate of mATG8s and will be valuable in decoding the biological functions of the individual LC3/GABARAPs.

Keywords: ATG8; LC3; immunofluorescence; phage display; selective autophagy.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Autophagy*
  • Autophagy-Related Protein 8 Family / analysis*
  • Biosensing Techniques / methods*
  • Cell Line
  • Fluorescence
  • Humans
  • Mitochondria / metabolism
  • Peptide Library
  • Protein Binding
  • Salmonella / immunology
  • Staining and Labeling / methods*


  • Autophagy-Related Protein 8 Family
  • Peptide Library