Vac8 spatially confines autophagosome formation at the vacuole in S. cerevisiae

ABSTRACT Autophagy is initiated by the formation of a phagophore assembly site (PAS), the precursor of autophagosomes. In mammals, autophagosome formation sites form throughout the cytosol in specialized subdomains of the endoplasmic reticulum (ER). In yeast, the PAS is also generated close to the ER, but always in the vicinity of the vacuole. How the PAS is anchored to the vacuole and the functional significance of this localization are unknown. Here, we investigated the role of the PAS–vacuole connection for bulk autophagy in the yeast Saccharomyces cerevisiae. We show that Vac8 constitutes a vacuolar tether that stably anchors the PAS to the vacuole throughout autophagosome biogenesis via the PAS component Atg13. S. cerevisiae lacking Vac8 show inefficient autophagosome–vacuole fusion, and form fewer and smaller autophagosomes that often localize away from the vacuole. Thus, the stable PAS–vacuole connection established by Vac8 creates a confined space for autophagosome biogenesis between the ER and the vacuole, and allows spatial coordination of autophagosome formation and autophagosome–vacuole fusion. These findings reveal that the spatial regulation of autophagosome formation at the vacuole is required for efficient bulk autophagy.