PDK

Serum-deprived HEK293-AT1A cells were pretreated with PD98059 (20 uM; 1 h) or OSU03012 (10 uM; 6 h) prior to 5 min stimulation with AngII (100 nm) or SII (50 uM). A, abundance of phospho-Akt T308 in whole cell detergent lysates. Representative phospho-Akt and total Akt immunoblots are shown above a bar graph depicting the mean ?S.E. of three biological replicates.

Induction of basal p62 Ser403 phosphorylation in ULK1/2-deficient MEF was suppressed by BX-795, an inhibitor of TBK1. WT ( Ulk1 +/+/Ulk2 +/+) and Ulk1 -/- /Ulk2 -/-(KO) MEF were treated with NaCl/Pi (Con) or 10 uM BX-795 for 6 h, and subjected to immunoblotting with indicated antibodies.

The inhibition of PI3K/Akt pathway is involved in ATG-induced FOXO3a dephosphorylation in PDGF-BB-activated HSCs. Serum-starved LX-2 human HSCs were pretreated with or without DMSO (vehicle) or ATG (0.5 uM) for 4 h, in the absence or presence of PI3K inhibitor LY294002 (20 uM) or Akt inhibitor PHT-427 (20 uM), and then incubated with or without 50 ng/ml PDGF-BB for 4 h. After treatment, aliquots of whole cell lysates were collected and subjected to Western blotting analysis. The experiments were repeated three times with similar results and a representative blot was shown for each protein. Total protein levels of Akt and FOXO3a served as internal controls.

(G) Effect of the PDK1 inhibitor GSK2334470 on MYC and P70 phosphorylation in CNE2/235 cell line by immunoblotting analysis. (H) Sensitivity to GSK2334470 in CNE2/235 cells after treatment with GSK2334470 for 5 d using the MTT assay. (I) Inhibitory effect of GSK2334470 in combination with BEZ235 on cell proliferation in CNE2/235 cells using the MTT assay. (J) IC50 values of BEZ235 with or without GSK2334470 in CNE2 and CNE2/235 cell lines. The data shown are representative of 3 individual experiments.

Serum-deprived HEK293-AT1A cells were pretreated with PD98059 (20 uM; 1 h) or OSU03012 (10 uM; 6 h) prior to 5 min stimulation with AngII (100 nm) or SII (50 uM). A, abundance of phospho-Akt T308 in whole cell detergent lysates. Representative phospho-Akt and total Akt immunoblots are shown above a bar graph depicting the mean ?S.E. of three biological replicates.

Induction of basal p62 Ser403 phosphorylation in ULK1/2-deficient MEF was suppressed by BX-795, an inhibitor of TBK1. WT ( Ulk1 +/+/Ulk2 +/+) and Ulk1 -/- /Ulk2 -/-(KO) MEF were treated with NaCl/Pi (Con) or 10 uM BX-795 for 6 h, and subjected to immunoblotting with indicated antibodies.

The inhibition of PI3K/Akt pathway is involved in ATG-induced FOXO3a dephosphorylation in PDGF-BB-activated HSCs. Serum-starved LX-2 human HSCs were pretreated with or without DMSO (vehicle) or ATG (0.5 uM) for 4 h, in the absence or presence of PI3K inhibitor LY294002 (20 uM) or Akt inhibitor PHT-427 (20 uM), and then incubated with or without 50 ng/ml PDGF-BB for 4 h. After treatment, aliquots of whole cell lysates were collected and subjected to Western blotting analysis. The experiments were repeated three times with similar results and a representative blot was shown for each protein. Total protein levels of Akt and FOXO3a served as internal controls.

(G) Effect of the PDK1 inhibitor GSK2334470 on MYC and P70 phosphorylation in CNE2/235 cell line by immunoblotting analysis. (H) Sensitivity to GSK2334470 in CNE2/235 cells after treatment with GSK2334470 for 5 d using the MTT assay. (I) Inhibitory effect of GSK2334470 in combination with BEZ235 on cell proliferation in CNE2/235 cells using the MTT assay. (J) IC50 values of BEZ235 with or without GSK2334470 in CNE2 and CNE2/235 cell lines. The data shown are representative of 3 individual experiments.