Fig. 5: ERG and β-Catenin co-occupy the PSG loci and co-regulate their expression.

a UCSC Genome Browser screenshots showing occupancy of ERG and β-Catenin in UMPS and RRM2 gene loci revealed by ERG ChIP-seq in VCaP cells and β-Catenin ChIP-seq (GSE53927). b,c ChIP-qPCR analysis of occupancy of ERG (b) and β-Catenin (c) at UMPS, RRM1, RRM2 and TYMS gene loci in VCaP cells. d,e Western blot (d) and RT-qPCR (e) analysis of indicated proteins and mRNAs in VCaP cells stably expressing indicated shRNAs. f Western blot analysis of indicated proteins in VCaP cells expressing indicated shRNAs. g Quantitative data showing the levels of UMP, dTDP and dTTP measured by LC-MS in VCaP cells generated as in (f). h MTS assay in VCaP cells with indicated shRNA as (f) and/or deoxynucleotides. i Western blot analysis of UMPS, RRM1 and RRM2 proteins in VCaP cells expressing indicated sgRNAs. j MTS assay in VCaP cells with depletion of indicated proteins as in (i). k–m VCaP cells expressing doxycycline (Dox)-inducible shRNAs for UMPS, RRM1 and RRM2 (sh3 PSGs) were inoculated into SCID mice and mice were treated with Dox daily for 30 days. Representative images of tumors harvested at the end of treatment (k), tumor growth curve (l) and tumor weight (m). Data in (b, c, e) and (g) were shown as mean ± s.d. from three independent experiments. The western blot assays in (d, f) and (i) were repeated two independent times with similar results. Data in (h) and (j) were shown as mean ± s.d. from five replicates. Data in (l) and (m) were shown as mean ± s.d. from six xenografts. Two-tailed Student’s t test was performed in (b, c, e, g) and (m). Two-way ANOVA was performed in (h, j) and (l).