a, Cross-linking of Atg18. Purified recombinant Atg18wt or Atg18FGGG (1.5 µM) was incubated with liposomes containing 5% PI3P, PI(3,5) P2, PI(4,5) P2 or 15% PS as a control for negative charge. After addition of a cleavable cross-linker or buffer only, liposomes were pelleted, dissolved in sample buffer, incubated in the presence or absence of 100 mM dithiothreitol (DTT) for 5 min at room temperature and analysed by SDS-PAGE and Coomassie staining. Cross-link products migrate at the upper limit of the separating gel. 