RNA-seq and ChIP-seq analysis of BMI1 or RING1B-silenced prostate cancer cells C4-2

Purpose: Study of the role of BMI1 dependent and indpendent of PRC1 in castration-resistant prostate cancer(CRPC) Method: The expression of BMI1 or RING1B was silenced by 2 independent siRNA strands targeted at BMI1 or RING1B in C4-2 cells, scramble RNA as control. mRNA profiles and genome-wide chromatin-state maps were generated by deep sequencing. AR, BMI1 and IgG ChIP was conducted in C4-2 cells. Results: AR-induced genes were significantly down regulated by BMI1 knockdown but not RING1B. higher expression levels of BMI1 activated genes (those downregulated by BMI1 knockdown) were significantly associated with poorer disease-free and poorer overall survival. Overall design: Expression profiling by RNA-seq, profiling of BMI1 and RNF2 (wildtype and knockdown) from C4-2 cell lines. DNA was purified from chromatin immunoprecipitates for AR, BMI1 or IgG using the phenol/chloroform extraction. IgG was used as a negative control for ChIP. Then, DNA was amplified by quantitative PCR and normalized to input.