Activated Atg15 is sufficient to disrupt ABs. (A) Schematic view of the in vitro assay employed to evaluate AB disruption. (B) Disruption of ABs by activated Atg15. Isolated ABs were incubated with 0.2% Tx, _prcAtg15, or _prcAtg15^S332A eluates at 30°C for 0 or 3 h. Eluate was added at two concentrations relative to standard assay conditions (1× and 3×). The reaction mixture was subjected to western blotting using α-Ape1 and α-GFP antibodies, respectively. (C) A model of AB disruption by active Atg15 in the vacuole. (i) Upon delivery to the vacuole, Atg15 is cleaved from MVB vesicles by Pep4/Prb1. (ii) Processed forms of Atg15 are tightly bound to the AB membrane. Atg15 is activated by Pep4/Prb1 during or following localization to the AB membrane. (iii) The active form of Atg15 hydrolyzes acyl ester linkages of phospholipids of AB membranes, releasing two acyl groups and disrupting the membrane. (iv) AB contents are released into the vacuole, where they are degraded by vacuolar hydrolases including Pep4 and Prb1. Source data are available for this figure: SourceData F6.