Fig. 2

GYS1 and glycogen are increased in hypoxia and acute GYS1 knockdown impairs breast cancer proliferation. a Protein expression of total GYS1, pGYS1(Ser641) (inactive form), the hypoxic markers HIF1 α and CA9, and other key enzymes in glycogen metabolism GBE1, PYGB and PYGL. Cells were cultured for 24 h in normoxia (N) or 1% hypoxia (H). b Glycogen levels after 24 h normoxia or 1% hypoxia, normalized by cell number (mean ± SD). c Breast cancer cell number after 5 days with siRNA-mediated GYS1 knockdown or scrambled control in normoxia (left) and 0.1% hypoxia (right) d Spheroid growth of MDA-MB-231 cells transfected with siRNA-mediated GYS1 knockdown or scrambled control. CA = carbonic anhydrase; GBE = glycogen branching enzyme; GYS = glycogen synthase; HIF = hypoxia-inducible factor; PYGB = brain glycogen phosphorylase; PYGL = liver glycogen phosphorylase; ns = not significant, *P < 0.05, **P < 0.01, ***P < 0.005, ****P < 0.001