Fig. 7

PFKFB4-NCOA3-FBP1 forms regulatory loop. A) Luciferase activity of NCOA3 in 786O cells with adenoviral (Av) PFKFB4 overexpression upon high (25 mM) and low (5 mM) glucose culture, two-way ANOVA; B) Q-PCR showing mRNA expression of target genes and 3 pentose phosphate pathway (PPP) genes upon knockdown of PFKFB4 or NCOA3 with shRNAs, one-way ANOVA; C) Reproduced from ChIP-Atlas, shown was heatmap ranked by binding score of NCOA3 from high to low in different model cells, each row representing one gene; D) Venn diagram showing PPP genes that could bound NCOA3 shown in ChIP-Atlas and E) heatmap showing their binding scores; F) Reproduced from the Cancer Genome Atlas (TCGA) clear-cell renal cell carcinoma (KIRC) dataset, shown were OncoPrint of PFKFB4 and FBP1 expression of z-score > 2 by RNA-seq (left) and FBP1 expression in patients with or without PFKFB4 overexpression, with patients with FBP1 overexpression excluded (right), Student’s t-test; G) Q-PCR showing FBP1 expression in 786O cells with PFKFB4 or NCOA3 knockdown (shRNA) under different oxygen status, two-way ANOVA; H) Cell count detected using CCK-8 in 786O cells with overexpression and knockdown of indicated genes, two-way ANOVA; I) Glucose uptake and lactate secretion measured in 786O cells in 786O cells with overexpression and knockdown of indicated genes, two-way ANOVA. (All in vitro assays performed in triplicates and at least 3 biological replicates; ns = not significant; *P < 0.05; **P < 0.01; ***P < 0.001; ****P < 0.0001)