Fig. 3

Enhanced carbohydrate metabolism upregulated EPHB2 in HNSCC. A. The mRNA (upper) and protein (lower) expression levels of EPHB2 in the normal epithelial cell line from nasopharynx (NP69) and larynx (HuLa-PC) and HNSCC cell lines. a-Tubulin was used as a loading control. B. The protein expression levels of EPHB2 in lymphatic metastatic HNSCC and non-lymphatic metastatic HNSCC. a-Tubulin was used as a loading control. C. qRT-PCR (upper) and western blot analysis (lower) of EPHB2 under different concentration of pyruvate with or without TM-1. a-Tubulin was used as a loading control. D. The source of nuclear acetyl-CoA. E. Nuclear acetyl-CoA levels in the indicated groups. F. The levels of histone H3 and H4 acetylation. G. The luciferase activities of EPHB2 reporter in the indicated groups. (H) qRT-PCR (upper) and western blot analysis (lower) of EPHB2 in the indicated groups. (I) Nuclear acetyl-CoA levels under pyruvate, glucose and 2-DG treatment. (J) Schematic (left) of the CRISPR CAPTURE approach following the western blot (right) to assess the levels of histone H3 and H4 acetylation in EPHB2 promoter in LECs under pyruvate, glucose and 2-DG treatment. K. ChIP assays examining the enrichment of P300, H327ac and Pol II on the EPHB2 promoter under pyruvate, glucose and 2-DG treatment. L. qRT-PCR (upper) and western blot analysis (lower) of EPHB2 under pyruvate, glucose and 2-DG treatment. NC, non-targeting negative control