Fig. 3.

GTPBP4 promotes aerobic glycolysis in HCC. (A) Venn diagram illustrating the proteins identified as GTPBP4 binding proteins from GTPBP4 OE PLC/PRF/5 and GTPBP4 OE SMMC-7721 cell groups. (B) Enrichment analysis and functional classification of GTPBP4 specific binding proteins using IPA software (left). The 25 most abundant proteins are listed (right). (C) Flux map and ¹³C labeled glucose tracing of glycolysis and the TCA cycle. (D) Quantification of intermediate metabolites in glycolysis and the TCA cycle by gas chromatography and mass spectrometry, Q-Exactive-Plus mass-spectrometer, and liquid chromatography coupled with tandem mass spectrometry for different metabolites. n = 6, unpaired 2-tailed t-test. (E) Quantification of isotopes derived from ¹³C₆-labeled glucose intermediates in glycolysis and the TCA cycle was performed using mass spectrometry analysis. M+3 indicates two carbon-labeled PEP, pyruvate, and lactate; M+2 indicates citrate, cis-aconitate, succinate, fumarate, and malate. n = 6, unpaired 2-tailed t-test. (F) The extracellular acidification rate was determined in control and GTPBP4 knockdown HCCLM3 cells. n = 6, one-way ANOVA. (G–H) Lactate production (G) and glucose consumption (H) were measured in HCC cells. Error bars represent the mean ± SD from six independent experiments. n = 6, unpaired 2-tailed t-test. *P < 0.05, **P < 0.01, ***P < 0.001. Abbreviations: IPA: ingenuity pathway analysis; TCA cycle: tricarboxylic acid cycle; ECAR: Extracellular acidification rate.