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. 2020 Jan 16;77(2):228–240.e7. doi: 10.1016/j.molcel.2019.10.016

Figure 4.

Figure 4

Mitotic Phosphorylation of Autophagy Regulators Is Independent of mTOR

(A) HCT116 cells were treated with 50 nM paclitaxel (16 h) and received DMSO, 2 μM RO-3306, 1 μM trametinib, or 1 μM AZD8055 2 h prior to lysis.

(B) HCT116 cells were treated with 50 nM paclitaxel (16 h) and received DMSO, 1 μM AZD8055, 250 nM Torin-1, or 1 μM PP242 2 h prior to lysis.

(C) HCT116 cells were treated with 50 nM paclitaxel for 6 h until approximately half of cells were rounded (indicative of mitosis). 2 h prior to lysis, cells were treated with either DMSO or AZD8055 (1 μM). Flasks then underwent mitotic shake-off to enrich interphase (I; adherent) and mitotic (M; detached) cells.

(D) HeLa cells stably expressing WT-TFEB-GFP or Δ30-TFEB-GFP were treated as indicated: starvation (2 h), refeed (complete medium; 1 h), 50 nM paclitaxel (16 h), or 1 μM AZD8055 (2 h). Western blots are from a single experiment representative of three independent experiments.