Fig. 5
AMPK activation is required for the HIF-1α-induced SSP gene expression in response to serine/glycine deprivation. A-C Indicated cells were cultured with or without S/G(-) media for the indicated periods of time. Intracellular ATP and AMP levels (A), NADPH and NADP+ levels (B), and ROS levels (C) were measured. NS, not significant. D Indicated cells were cultured with or without S/G(-) media in the presence or absence of 2-DG (10 mM) for the indicated periods of time. Immunoblotting analyses were performed with the indicated antibodies. E Indicated cells were cultured with or without S/G(-) media for the indicated periods of time in the presence or absence of NAC (2 mM). Immunoblotting analyses were performed with the indicated antibodies. F Indicated cells were cultured with S/G(-) media for 24 h in the presence or absence of compound C (5 μM), and then treated with CHX (100 μg⋅mL−1) for the indicated periods of time. Immunoblotting analyses were performed with the indicated antibodies. Quantification of HIF-1α levels relative to tubulin is shown. Data represent the mean ± s.d. of three independent experiments. G Indicated cells with or without the depletion of AMPK were cultured with or without S/G(-) media for 24 h. Immunoblotting analyses were performed with the indicated antibodies. H HRE luciferase activities were measured in the indicated cells with or without the depletion of AMPK culturing with or without S/G(-) media for the indicated periods of time. NS, not significant. I and J Indicated cells with or without the depletion of AMPK were cultured with or without S/G(-) media for 24 h. Quantitative real-time PCR (I) and immunoblotting (J) analyses were performed with the indicated primers and antibodies, respectively. K Indicated cells were cultured with S/G(-) media in the presence or absence of compound C (10 μM) for 24 h. Quantitative real-time PCR analyses were performed with the indicated primers. The data represent the mean ± s.d. of three independent experiments (A-C, F, H, I, K). *P < 0.05; **P < 0.01; ***P < 0.001, based on the Student’s t-test