The carboxy terminus of yeast Atg13 binds phospholipid membrane via motifs that overlap with the Vac8-interacting domain

Macroautophagy/autophagy is a conserved catabolic recycling pathway involving the sequestration of cytoplasmic components within double-membrane vesicles termed autophagosomes. The autophagy-related (Atg) protein Atg13 is a key member of the autophagy initiation complex. The Atg13 C terminus is an intrinsically disordered region (IDR) harboring a binding site for the vacuolar membrane protein Vac8. Recent reports suggest Atg13 acts as a hub to assemble the initiation complex, and also participates in membrane recognition. Here we show that the Atg13 C terminus directly binds to lipid membranes via electrostatic interactions between positively charged residues in Atg13 and negatively charged phospholipids as well as a hydrophobic insertion of a Phe residue. We identified 2 sets of residues in the Atg13 IDR that affect its phospholipid-binding properties; these residues overlap with the Vac8-binding domain of Atg13. Our data indicate that Atg13 binding to phospholipids and Vac8 is mutually exclusive, and both are required for efficient autophagy.

Atg: autophagy-related; CD: circular dichroism; Cvt: cytoplasm-to-vacuole targeting; IDR: intrinsically disordered region; ITC: isothermal calorimetry; MIM: MIT-interacting motif; MKO: multiple-knockout; PAS: phagophore assembly site; PC: phosphatidylcholine; PS: phosphatidylserine; PtdIns: phosphatidylinositol; PtdIns3P: phosphatidylinositol-3-phosphate