Microautophagy is a mode of autophagy in which the lysosomal or vacuolar membrane invaginates and engulfs target components. Oku et al. show that, upon a diauxic shift, yeast microautophagy involves recruitment of ESCRT proteins to the vacuolar membrane, including clathrin-interacting Vps27, and uptake of lipid droplets by the vacuole.

No abstract

Autophagy is a lysosomal/vacuolar delivery system that isolates and degrades cytoplasmic material. Following delivery by autophagosomes, cytoplasmic components are released into the vacuole within an autophagic body (AB), which is a single-membrane structure derived from the inner membrane of the autophagosome. This membrane must be disrupted for degradation of the cytoplasmic cargo to occur. The vacuolar proteases Pep4 and Prb1, as well as the lipase Atg15, are known to be necessary for this process, but the mechanistic underpinnings remain unclear. In this study, we establish a system to detect lipase activity in the vacuole and use it to show that Atg15 is the sole vacuolar phospholipase and that Pep4 and Prb1 are required for the activation of Atg15 lipase function, which occurs following delivery of Atg15 to the vacuole by the MVB pathway. In vitro experiments also reveal that Atg15 is a B-type phospholipase of broad substrate specificity that is likely implicated in the disruption of a range of membranes delivered to the vacuole. Further, we use isolated ABs to demonstrate that Atg15 alone is able to disrupt AB membranes.

Autophagy is a major cellular degradation pathway that is highly conserved among eukaryotes. The identification of cargos captured by autophagosomes is critical to our understanding of the physiological significance of autophagy in cells. In the yeast S. cerevisiae, cells deficient in the vacuolar lipase Atg15 accumulate autophagic bodies (ABs) within the vacuole following the induction of autophagy. As ABs contain cytosolic components including proteins, RNAs, and lipids, their purification allows the identification of material targeted by autophagy for degradation. In this study, we demonstrate a method to purify intact ABs from vacuoles that retain membrane integrity and contain autophagic cargos. This technique offers a valuable tool for the identification of the cargos of autophagy, examination of autophagic cargo selectivity, and biochemical characterization of autophagosome membranes.