Fig 1: Glycolysis is hyperactivated in erythroblasts expressing only GATA1s. (A) Schematic representation of the metabolic fate of uniformly labeled 13C6-glucose through glycolysis and the TCA cycle. (B) 13C6-glucose tracing of glycolytic and TCA cycle metabolites (as shown in panel A) in 24-hour differentiated HUDEP1 cells expressing either GATA1FL or GATA1s (1D5). The ratio of labeled PEP isotopologs is shown on the right. (C) Intracellular PK activity measured in 24-hour differentiated HUDEP1 cells expressing GATA1FL or GATA1s (1D5 and 1KI9) using the Abcam PK Assay Kit (ab83432). (D) Extracellular lactate levels measured from culture media of 24-hour differentiated HUDEP1 cells expressing GATA1FL or GATA1s (1D5 and 1KI9), using the Abcam L-Lactate Assay Kit (ab65330). In panels B-D, asterisks indicate statistically significant differences. (E) CUT&RUN heat maps comparing H3K27la and H3K18la histone modifications in HUDEP1 cells expressing GATA1s (1D5) vs GATA1FL after 24 hours of differentiation. (F) IGV snapshots of CUT&RUN tracks for H3K27ac, H3K27la, and H3K18la at the PKM locus in HUDEP1s (1D5) vs GATA1FL cells. (G) Culture media lactate concentration in FL1 and 1D5 HUDEP1 clones treated with vehicle (DMSO) or GSK2837808A. (H) Western blot of lactylated lysine in extracts from GATA1FL1 and G1s_1D5 clones treated with DMSO or GSK2837808A. (I) Flow cytometry plots of erythroid differentiation in GATA1s_1D5 clone treated with varying doses of GSK2837808A at days 3 and 5. ∗P < .05; ∗∗P < .01. Acetyl-CoA, acetyl coenzyme A; diff, differentiation; GSK, GSK2837808A; ns, no statistical significance; Pyr, pyruvate; Suc, succinate; TCA, trichloroacetic acid; TES, transcription end site; TSS, transcription start site.