3.

Hexokinase (HK) II decreases relative activity of pyruvate dehydrogenase (PDH) complex and regulates metabolic flux. (A) Relative activity of PDH complex and P-S293 levels of endogenous PDHA1 were determined and quantified in HKII-overexpressed cells; (B) Relative activity of PDH complex and P-S293 levels of endogenous PDHA1 were determined and quantified after HKII knockdown; (C) PDK1 was knocked down in HeLa cells and then HKII was transfected into the cells, the activity of PDH complex was measured by PDH enzyme microplate assay kit (ab109902); (D) HKII-overexpressing HeLa cells were incubated under normoxia or hypoxia and the amount of glucose 6-phosphate (G6P), pyruvate, lactate, and citrate were determined and quantified. **, P<0.01; ***, P<0.001.