Figure 2.

HYlight does not affect fructose 1,6-bisphosphate (FBP) levels in liver cells, and its response is linked to glycolysis

(A) ECAR of HepG2 liver cancer cells after 1 h of glucose starvation and following the addition of 10 mM glucose, 10 μM oligomycin, and 100 mM 2-DG in two steps. The ECAR readout is in milli-pH units per min and normalized to cell number. Sample size is n = 3.

(B) OCR assay of HepG2 liver cancer cells after 1 h of glucose starvation and following the addition of 10 mM glucose, 10 μM oligomycin, and 100 mM 2DG in two steps. The OCR is in picomoles of oxygen per min and normalized to cell number. Sample size is n = 3.

(C) Fluorescence ratiometric images of HepG2 cells under 15 mM glucose with or without gluconeogenic stimulation. Scale bar: 200 μm.

(D) FBP generation is catalyzed by phosphofructokinase 1 (PFK1) in glycolysis and its destruction is mediated by fructose 1,6-bisphosphatase 1 (FBP1) in gluconeogenesis. Below, changes in the fluorescence ratio of HYlight in HepG2 liver cancer cells after 1 h of glucose starvation and sequential addition of 50 μM dexamethasone, 100 nM glucagon, and 15 mM glucose. Comparison between gluconeogenic medium (GNG) and normal media in the presence or absence of either 20 μM FBP1 inhibitor (top) or 20 μM PFK inhibitor (bottom). Data are presented as mean ± SEM. n = 3, with approximately 50 cells per condition and replicate. See also Figure S2 for additional details. Figure 2D can also be accessed through BioRender: BioRender.com/e27y335.