Mechanistic investigation of aberrantly expressed HOX genes by ChIP and qRT-PCR. (a) Differential expression of HOXA11 in VM-CUB1 and 5637 HOTAIR clones was verified by qRT-PCR relative to TBP (left graph). ChIP with subsequent qPCR was used to determine changes in H3K4, H3K9 and H3K27 trimethylation, each as compared to IgG control for VM-CUB1 cells (middle) and 5637 cells (right). Analogous analyses for HOXB2
(c) and HOXD10
(d). Overexpression of polycomb group proteins EZH2 (e, 85 kDa) and SUZ12 (f, 83 kDa) on the RNA (left) and protein level (right) in UC cells. Tubulin protein (50 kDa) was detected as a loading control.