Fig. 2

Inhibition of TOR reduces the acetylome through Gcn5. a The level of H3ac in histone acetyltransferases (HATs) disruption mutants. b Quantification of the relative levels of H3ac/H3 in (a). c Effects of GCN5 deletion, rapamycin (Rapa) treatment, and nitrogen starvation (MM-N) on histone acetylation. Total proteins extracted from mycelia of tested strains with or without treatment were immunoblotted using indicated antibodies. d Transcription of GCN5 detected by RT-PCR in the mycelia of PH-1 grown in completed medium (CM) and under autophagy induction condition (MM-N). e CM and rapamycin/nitrogen starvation–treated Gcn5-GFP-expressing mycelia were observed with a confocal microscope; intensity of Gcn5-GFP was quantified with ImageJ. Bar = 10 μm. f, g Relative amount of Gcn5-GFP under tested conditions in (e). Samples were analyzed using immunoblot assays. Data are presented as mean ± s.d from triplicates. Asterisks indicate significant difference according to a LSD test at P< 0.01. h Relative amount of Gcn5-GFP under CM, MM-N, or CM with rapamycin (25 nM) in the presence of the translational inhibitor, cycloheximide (CHX). i MG132 or 3-MA was added into nitrogen-free minimal or CM with (out) rapamycin medium for 2 h, then mycelia were harvested and lysed and the degradation of Gcn5-GFP was detected with immunoblotting. j Degradation of Gcn5-GFP in the 26S proteasome–defective mutant Δ26S-RS10, or the autophagy of defective mutant Δatg8. k The ubiquitination level of Gcn5-Flag under autophagy induction conditions