Discovery of pan autophagy inhibitors through a high-throughput screen highlights macroautophagy as an evolutionarily conserved process across 3 eukaryotic kingdoms

Autophagy. 2017 Sep 2;13(9):1556-1572. doi: 10.1080/15548627.2017.1339002. Epub 2017 Aug 9.


Due to the involvement of macroautophagy/autophagy in different pathophysiological conditions such as infections, neurodegeneration and cancer, identification of novel small molecules that modulate the process is of current research and clinical interest. In this work, we developed a luciferase-based sensitive and robust kinetic high-throughput screen (HTS) of small molecules that modulate autophagic degradation of peroxisomes in the budding yeast Saccharomyces cerevisiae. Being a pathway-specific rather than a target-driven assay, we identified small molecule modulators that acted at key steps of autophagic flux. Two of the inhibitors, Bay11 and ZPCK, obtained from the screen were further characterized using secondary assays in yeast. Bay11 inhibited autophagy at a step before fusion with the vacuole whereas ZPCK inhibited the cargo degradation inside the vacuole. Furthermore, we demonstrated that these molecules altered the process of autophagy in mammalian cells as well. Strikingly, these molecules also modulated autophagic flux in a novel model plant, Aponogeton madagascariensis. Thus, using small molecule modulators identified by using a newly developed HTS autophagy assay, our results support that macroautophagy is a conserved process across fungal, animal and plant kingdoms.

Keywords: autophagic flux; autophagy; high-throughput screening; kingdoms; peroxisome; programmed cell death; small molecules.

MeSH terms

  • Amino Acid Chloromethyl Ketones / pharmacology
  • Animals
  • Autophagosomes / drug effects
  • Autophagosomes / metabolism
  • Autophagy* / drug effects
  • Biological Evolution*
  • Embryo, Mammalian / cytology
  • Enzyme Assays
  • Eukaryotic Cells / drug effects
  • Eukaryotic Cells / metabolism*
  • Fibroblasts / drug effects
  • Fibroblasts / metabolism
  • HeLa Cells
  • High-Throughput Screening Assays / methods*
  • Humans
  • Luciferases / metabolism
  • Magnoliopsida / drug effects
  • Mice
  • Models, Biological
  • Nitriles / pharmacology
  • Saccharomyces cerevisiae / drug effects
  • Saccharomyces cerevisiae / metabolism
  • Small Molecule Libraries / pharmacology
  • Sulfones / pharmacology


  • 3-(4-methylphenylsulfonyl)-2-propenenitrile
  • Amino Acid Chloromethyl Ketones
  • Nitriles
  • Small Molecule Libraries
  • Sulfones
  • ZPCK
  • Luciferases