A6358

Anti-Vps34 (1:1,000, no. 4263), Beclin 1 (1:1,000, no. 3738), acetylated lysine (1:500, no. 9441), haemagglutinin (HA)-Tag (1:1,000, no. 3724, Rabbit) and HA-Tag (1:1,000, no. 2367, Mouse) antibodies were purchased from Cell Signaling Technology. Anti-UVRAG (1:1,000, NBP1–18885), Beclin 1 (1:1,000, NBP1–00088) and LC3 (1:5,000, NB100–2220) antibodies were obtained from NOVUS. Anti-Rubicon (1:1,000, ab92388) and p150 (1:1,000, ab128903) antibodies were obtained from Abcam. Anti-SIRT1 (1:1,000, AJ1717a), SIRT2 (1:1,000, AJ1718a) and HDAC1 (1:1,000, AP1101a) antibodies were purchased from Abgent. Anti-p300 (1:500, sc-585), p62/SQSTM1 (1:2,000, sc-28359) and HDAC2 (1:1,000, sc-55542) antibodies were purchased from Santa Cruz Biotechnology. Anti-a-tubulin (1:2,000, T6199), Flag (1:5,000, F3165) and Atg14L (1:1,000, A6358) antibodies were obtained from Sigma. Anti-phospho S409 Beclin 1 antibodies were generated by immunizing rabbits with the corresponding phosphopeptides. Bafilomycin A1, rapamycin, deacetylase inhibitors TSA and NAM, and CK1 inhibitor D4476 were purchased from Sigma.

The following antibodies were used: mouse monoclonal anti-NLRX1 (1:1000; 04–146; Millipore), rabbit polyclonal anti-VPS34/PI3KC3 (38–2100; Invitrogen), rabbit polyclonal anti-Beclin 1 (1:1000; 3738; Cell Signaling Technology), rabbit polyclonal anti-ATG14 (1:1000; A6358; Sigma-Aldrich), rabbit polyclonal anti-UVRAG (1:1000; NBP1-18885; Novus Biologicals), rabbit polyclonal anti-Rubicon (1:1000; ab92388; Abcam), mouse monoclonal anti-LAMP1 (1:100; H4A3; Santa Cruz Biotechnology, Santa Cruz, CA), mouse anti-polyubiquitin (FK2) (1:100; Nippon Bio-Test Laboratories), rabbit monoclonal anti-α-tubulin (1:1000; T6199; Sigma-Aldrich), mouse monoclonal anti-FLAG M2 (1:1000; F1804; Sigma-Aldrich), and mouse monoclonal anti-GFP (1:1000; GF200; 04363-24; Nacalai Tesque). Horseradish peroxidase-conjugated anti-mouse or anti-rabbit IgG (1:5000; Jackson ImmunoResearch Laboratories) were used as secondary antibodies for immunoblotting. The fluorescent secondary antibodies Alexa Fluor 488-conjugated goat anti-rabbit IgG (1:200; Molecular Probes/Invitrogen), or Alexa Fluor 568-conjugated goat anti-mouse IgG (1:200; Molecular Probes/Invitrogen) were used for immunofluorescence.