Chloroquine

Nutlin-3a and Erlotinib were purchased from Selleck chemicals (Houston, TX, USA). MG132 was from Beyotime Biotechnology (Shanghai, China). Chloroquine was from Abcam (Cambridge, UK), and all other chemicals were of analytical grade and purchased from Sigma Chemical (Burlington, USA). For in vitro and in vivo studies, Nutlin-3a was reconstituted in dimethyl sulfoxide (DMSO) and 2% DMSO/1% hydroxyethyl cellulose/0.2% Tween-80, respectively.

NDGA (Sigma), C646 (Sigma), A485 (Tocris), and Bafilomycin A1 (Adipogen) were dissolved in dimethyl sulfoxide (DMSO) (Sigma-Aldrich), Chloroquine (Biovision) was dissolved in water. Phosphate-buffered saline (PBS) was purchased from Corning. Tris-buffered saline with Tween (TBST) buffer (150 mM NaCl, 0.01% (v/v) Tween-20, 50 mM Tris-HCl buffer, pH 7.6) was used for immunoblotting. Blocking buffer and primary antibody incubation solution was 5% (w/v) BSA (Sigma), pH 7.0, in TBST. Secondary antibody incubation buffer was 5% (w/v) blocking reagent (Sigma) in TBST.

Worms were synchronized by treating gravid adults with bleaching solution (H₂O:bleach:5 N NaOH 7:2:1). Animals were washed three times in M9 buffer and incubated in M9 buffer on a rotor. On days 5 and 10, 30 μl of M9 were plated on an NGM plate; after 1 h, the percentage of alive larval stage 1 worms was measured. For the colocalization assays, starvation was performed for 6 h and chloroquine (catalog no. 1825; BioVision) was added on plates at 5 μM concentration 3 h before imaging. For heat stress, day 1 adult animals were placed in a 37 °C incubator for 1 h and survival was scored on days 2 and 4 of adulthood. For ultraviolet-C-induced DNA damage, day 1 worms were placed on ultraviolet-inactivated OP50 bacteria and treated with either 200 J m⁻² or 400 J m⁻² ultraviolet-C irradiation using an ultraviolet crosslinker (BIO-LINK-BLX-E365; Vilber Lourmat).