Fig. 4

PKM2 activation reduces ATP pools leading to AMPK activation and energy crisis. a Cellular ATP levels in response to 72 h DASA-58 (15 μM) treatment in different BCa cell lines showing slight reduction in ATP levels in MCF7, MDA MB 231, and HCC1443 cell lines. b Western blot analysis of different BCa cell lines in response to 72 h treatment of either DASA-58 (15 μM) or TEPP-46 (30 μM), showing levels of pAMPK (T172) and total AMPK. c Western blot showing the induction in pAMPK (T172) levels in MCF7 cells in response to 72 h DASA-58 (15 μM) treatment in triplicates (upper panel), with the corresponding densitometric analysis representing pAMPK/tAMPK as fold change normalized to actin levels, and the induction of pAMPK (T172) levels in MCF7 cells in response to DASA-58 (15 μM) at 24, 48, and 72 h of treatment (lower panel). d Western blot analysis of different BCa cell lines showing the expression of pACC, tACC, and vinculin in response to either DASA/58 (15 μM) or TEPP-46 (30 μM); numbers underneath the figure represent the densitometric analysis of pACC/tACC band intensities normalized to the corresponding vinculin bands. e Western blot analysis showing the levels of pAMPK/tAMPK and of pACC/tACC together with vinculin in response to increasing concentrations of DASA-58 and TEPP-46 in MCF7 and MDA MB 231. f Relative ATP levels in MCF7 in response to DASA-58 (15 μM) alone or combined with oligomycin (5 μM) or 2DG (5 mM). g, h Western blots with pAMPK (T172) and TXNIP levels in response to either DASA-58 (15 μM) alone or combined with mitochondrial modulators (g) or glycolytic branching/related pathways modulators (h) showing that none of the used combo treatments is able to rescue TXNIP levels in the presence of DASA-58 and that mitochondrial inhibitors enhance PKM2 effects on activating AMPK signaling (depicted as T172 phosphorylation of AMPK)