Reversible One-way Lipid Transfer at ER-Autophagosome Membrane Contact Sites via Bridge-like Lipid Transfer Protein Atg2

Bridge-like lipid transfer proteins (LTPs) contain a repeating β-groove domain and long hydrophobic grooves that act as bridges at membrane contact sites (MCSs) to efficiently transfer lipids. Atg2 is one such bridge-like LTP essential for autophagosome formation, during which a newly synthesized isolation membrane (IM) emerges and expands through lipid supply. However, studies on Atg2-mediated lipid transfer are limited to in vitro studies due to the lack of a suitable probe for monitoring phospholipid dynamics in vivo . Here, we characterized the lipophilic dye octadecyl rhodamine B (R18), which reached the endoplasmic reticulum (ER) through the actions of flippases and oxysterol-binding protein-related proteins. Using R18, we demonstrated phospholipid transfer from the ER to the IM during autophagy in vivo . Upon autophagy termination, we observed reversible phospholipid flow from the IM to the ER in response to environmental changes. Our findings highlight the critical role of bridge-like LTPs in MCS-mediated phospholipid homeostasis.