Fig. 6

ULK phosphorylates β1-Ser108 in cells. Statistical analyses were performed using one-way ANOVA with post hoc Dunnett’s multiple comparison test, unless indicated. Immunoblots for β1-pSer108 and α-pThr172 in KI-α1β1γ1 purified from HEK293T cells incubated with a 1 mM H₂O₂ and 10 μM 6965 for 45 min, or b 2 mM phenformin and 10 μM 6965 for 1 h. n = 3, representative immunoblots shown. Error bars, mean % phosphorylation relative to H₂O₂- or phenformin-treated ± s.e.m. **P < 0.01 indicates significant increase, and ^## P < 0.01, ^### P < 0.001 and ^#### P< 0.0001 indicate significant decrease, compared to H₂O₂- or phenformin-treated. c Immunoblots for β1-pSer108 and α-pThr172 in lysates from HEK293T cells, WT or ulk1/2-dKO iMEFs incubated in 25 mM glucose DMEM + 10% serum. n = 3 individual cultures per cell line, representative immunoblots shown. Error bars, mean fold increase in phosphorylation relative to HEK293T cells ± s.e.m. ****P < 0.0001 indicates significant increase in phosphorylation compared to HEK293T cells. d Immunoblots for β1-pSer108 and α-pThr172 in KI-α1 AMPK purified from WT or ulk1/2-dKO iMEFs stimulated with 2 mM phenformin for 1 h. n = 3, representative immunoblots shown. Error bars, mean fold increase in phosphorylation relative to basal ± s.e.m. Statistical analysis was performed using unpaired two-tailed Student’s t-test. *P < 0.05 indicates significant decrease compared to WT iMEFs