Fig. 7: FBP and its analogues regulate PGAM1 activity through histidine phosphorylation in living cells.

a PGAM1 histidine phosphorylation in living HEK293T cells with overexpressed flag-tagged PGAM1 treated by FBP (10 mM) for 30 min in glucose-free medium. b Intracellular FBP concentration and 2-PG/3-PG ratio in HepG2 cells treated by FBP (10 mM) for 30 min in glucose-free medium. c Lactate production and glucose consumption in sgCTRL or sgPGAM1 HepG2 cells treated by FBP (10 mM) for 30 min in 10 mM glucose medium. d Cell proliferation of sgCTRL or sgPGAM1 HepG2 cells treated by FBP (10 mM) for 24 h in glucose-free medium. e Cell viability of HepG2 cells treated by FBP analogues 1-DMeFBP, 6-DMeFBP and 1,6-DMeFBP (1 mM) for 24 h in high glucose (25 mM) medium respectively. f PGAM1 histidine phosphorylation in living HepG2 cells with overexpressed flag-tagged PGAM1 treated by 1-DMeFBP (1 mM) for 24 h. g Lactate production and glucose consumption in sgCTRL or sgPGAM1 HepG2 cells treated by 1-DMeFBP (1 mM) for 24 h in high glucose medium. h Cell proliferation of sgCTRL or sgPGAM1 HepG2 cells treated by 1-DMeFBP (1 mM) for 24 h in high glucose medium. (i) Schematic model depicting the intrinsic intrapathway positive feedback regulation of glycolysis by FBP-PGAM1 covalent signaling or FBP analogue-PGAM1 inhibition. All measurements are presented as mean ± SD for three biological replicates. Statistical differences were determined by one-way ANOVA followed by Dunnett’s multiple comparison tests (a, b, e, f) or two-sided Student’s t-test (c, d, g, h). Source data are provided as a Source Data file.