Fig. 6

Atg8 is partially deacetylated by the deacetylase Hdf1. a Co-immunoprecipitation of GFP-Atg8 and mCherry-Hdf1. b In vitro pull-down of GST-Hdf1 and GFP-Atg8. Purified GST-Hdf1 was incubated with the lysate of the fungal strain expressing GFP-Atg8. GFP-Atg8 was immunoprecipitated with an anti-GFP agarose, the precipitated proteins were detected with anti-GFP and anti-GST antibodies. c, d Autophagy process in WT and Δhdf1 mutant. c Translocation of GFP-Atg8 in mycelia of PH-1 and Δhdf1 under starvation. Vacuoles were stained with the CMAC dye. Bar = 10 μm. d The cleavage of GFP-Atg8 in PH-1 and Δhdf1 under starvation. e Acetylation of GFP-Atg8 in PH-1 or Δhdf1. GFP-Atg8 was immunoprecipitated from mycelial lysate of PH-1 or Δhdf1 under CM or MM-N. Acetylation level of GFP-Atg8 was detected with the anti-acetyl-lysine (α-AcK) antibody. f The amount of Atg8 protein in nuclear fraction of PH-1 and Δhdf1 under CM or MM-N. Nuclear fraction was prepared and subjected to immunoblot assay with the anti-GFP antibody. Hybridization with anti-H3 antibody served as an internal control of nuclear protein sample