Figure 2.

The yeast vacuole acts as a platform for PAS assembly and phagophore anchoring. A. The vacuolar protein Vac8 mediates the contact with the Phagophore Assembly Site (PAS) machinery via its interaction with Atg13 or Atg11. Vac8 is also involved in the formation of the vacuole MCS with the nucleus (N). Atg9 vesicles are docked at the PAS through the Atg1 complex and/or Atg11 and represent the nucleation point for phagophore formation. COPII vesicles also contribute to phagophore growth. Specific autophagy proteins gather at the phagophore–vacuole MCS to form the “VICS cluster”. At this stage, Atg8 is conjugated to the phagophore membrane, while Atg9 localizes to the rim, where the ER–phagophore MCS is established. During maturation, Atg8 is removed from the outer membrane and the autophagosome is thought to lose contact with the vacuole. Finally, the fusion machinery mediates tethering to the vacuole and membrane fusion, leading to subsequent degradation of the autophagosome. B. The strong tethering of the phagophore to the vacuole at the MCS is shown in tomogram slices of in situ correlative cryo-ET. Two different morphologies of the contact site are observed: peak (1) or extended (2). Scale bars 100 nm (Adapted from Bieber et al., 2022; left tomogram: EMD-15549).