FIG 10.

Assay for Atg8 lipidation. To determine the requirements of phosphorylation sites for Atg8 lipidation, BbATG3 and its mutants were transformed into the ΔBbatg3 mutant strain with the GFP-tagged ATG8. For aerial and submerged mycelia, fungal strains were cultured on SDAY for 5 days and in SDB for 2 days, respectively. For starvation stress, the submerged mycelia were incubated in mineral solution for 1, 3, and 6 h. The cell lysate was resolved on an SDS-PAGE gel with urea, and GFP-Atg8 was probed with anti-GFP antibody. Protein quality in the sample was controlled by actin. Arrowheads and asterisks indicate the GFP-Atg8 and its phosphatidylethanolamine-tailed form. “+” and “−” indicate presence and absence of the indicated genetic factor.