Calcium transients on the ER surface trigger liquid-liquid phase separation of FIP200 to specify autophagosome initiation sites

Cell. 2022 Oct 27;185(22):4082-4098.e22. doi: 10.1016/j.cell.2022.09.001. Epub 2022 Oct 4.

Abstract

The mechanism that initiates autophagosome formation on the ER in multicellular organisms is elusive. Here, we showed that autophagy stimuli trigger Ca2+ transients on the outer surface of the ER membrane, whose amplitude, frequency, and duration are controlled by the metazoan-specific ER transmembrane autophagy protein EPG-4/EI24. Persistent Ca2+ transients/oscillations on the cytosolic ER surface in EI24-depleted cells cause accumulation of FIP200 autophagosome initiation complexes on the ER. This defect is suppressed by attenuating ER Ca2+ transients. Multi-modal SIM analysis revealed that Ca2+ transients on the ER trigger the formation of dynamic and fusion-prone liquid-like FIP200 puncta. Starvation-induced Ca2+ transients on lysosomes also induce FIP200 puncta that further move to the ER. Multiple FIP200 puncta on the ER, whose association depends on the ER proteins VAPA/B and ATL2/3, assemble into autophagosome formation sites. Thus, Ca2+ transients are crucial for triggering phase separation of FIP200 to specify autophagosome initiation sites in metazoans.

Keywords: ATG9; Ca(2+) transient; EI24; ER; FIP200; autophagosome formation; liquid-liquid phase separation; lysosome.

MeSH terms

  • Animals
  • Autophagosomes* / metabolism
  • Autophagy
  • Autophagy-Related Proteins / metabolism
  • Calcium* / metabolism
  • Cell Cycle Proteins / metabolism
  • Endoplasmic Reticulum / metabolism

Substances

  • Calcium
  • Autophagy-Related Proteins
  • Cell Cycle Proteins