PFKP is a direct onco-target of YY1 in prostate cancer. (A, B) Volcano plots of the RNA-seq profiles highlight PFKP among the most altered transcripts upon YY1 depletion in 22Rv1 (A) and C4-2 (B) cells. (C) Correlation between the expression levels of PFKP and YY1 among prostate cancer patients using the StandUp2Cancer (SU2C) dataset (43). (D, E) Box plots showing the PFKP expression levels among patient samples from the indicated cohort reported by Tomlins et al (30) (D) or Varambally et al (42) (E). (F) Immunoblotting of endogenous YY1 and PFKP post-KD of YY1, relative to mock, in 22Rv1 (left) and C4-2 (right) cells. (G) Immunoblotting of exogenous YY1 (using HA antibody) and endogenous PFKP post-transduction of HA-tagged YY1^WT or YY1^S365D into 22Rv1 (left) or C4-2 (right) cells with stable YY1 KD (by shYY1#94, which targets 3'-UTR of YY1). (H) IGV views of the YY1 ChIP-seq profile at the proximal promoter of PFKP. (I, J) Relative transcription activities from a luciferase reporter that carries the PFKP promoter (right panel), either –2008 to +7 (I) or –575 to +37 bp (J) from transcription start site, after its co-transduction with HA-tagged YY1^WT or YY1^S365D into 22Rv1 cells. Left panels show the reporter results using the cells transduced with the empty luciferase reporter. Y-axis shows the averaged fold-change in the reporter signals after normalization to those of internal luciferase control and then to mock samples (the first sample); n = 3 independent experiments; ** P< 0.01. (K) Scheme shows the used mutation (mut) of putative YY1-binding sites within the PFKP promoter. Empty box denotes the mutated YY1-binding site. (L, M) Luciferase activity from a PFKP promoter reporter, which carries either WT or the indicated mutation of YY1-binding sites (refer to K), after its co-transduction with WT YY1 cDNA into 22Rv1 (L) or C4-2 cells (M). Y-axis shows the averaged fold-change in the reporter signals after normalization to those of internal luciferase controls and then to empty vector-transfected cells (n = 3 independent experiments). * P < 0.05; ** P < 0.01; *** P < 0.001; NS, not significant.