Transcription is regulated through a variety of different mechanisms. One facet of transcriptional regulation is epigenetics, which is the regulation of gene expression without altering the nucleotide sequence of the genome, but rather reversible modifications, e.g. CpG methylation. The CpG islands in tumor suppressor gene promoters are often hypermethylated, thus down regulating their expression. The p73 gene is a member of the p53 tumor suppressor family of proteins, and like p53, has a bifurcated promoter. The p73 P1 promoter transcribes a full-length mRNA, which then translates into the transcriptionally active p73. The p73 P2 promoter transcribes a truncated mRNA, which translated into transcriptionally inactive ΔNp73. The ΔNp73 product lacks the N-terminal trans-activation domain that TAp73 possesses. This study sought to analyze the extent of DNA methylation in the CpG islands in the p73 P1 promoter of three prostate cancer (PCa) cell lines (LNCaP, PC-3, and DU145), and primary prostate cells. We hypothesized that the p73 P1 promoter would be hypermethylated in prostate cancer cell lines and hypomethylated in primary prostate cells. The methylation status of the p73 P1 promoter in all four prostate cell lines was analyzed by using bisulfite sequencing PCR (BSP). Our analyses revealed that a CpG island in the p73 P1 was hypermethylated in LNCaP, PC-3, and DU145 cells compared to primary prostate cells. Furthermore, among the three PCa cell lines evaluated, the p73 P1 promoter was the most methylated in DU145 cells, and the least methylated in LNCaP cells. The P1 region sequenced contained 28 CpG islands, and among them, only five were methylated in the primary prostate cells (18% methylated), 23 were methylated in LNCaP cells (82% methylated), 26 in PC-3 cells (93% methylated), and 28 in DU145 cells (100% methylated). Furthermore, of the 28 CpG sites, 14 of them were differentially methylated in the primary prostate cell line versus the three PCa cell lines evaluated in this study. These data are consistent with the PCa tumor-derived cell lines (LNCaP, PC-3, and DU145) having hypermethylated CpG sites in the p73 P1 promoter compared to the primary prostate cell line.

Citation Format: Nicholas E. Braganca, L. Michael Carastro, Johannes J. Schabort, Jong Y. Park. Bisulfite DNA sequencing analyses to detect methylation patterns in the p73 gene promoter in prostate cancer cell lines [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 4363. doi:10.1158/1538-7445.AM2017-4363