Abstract 468: Cancer cells with G/G mdm2 SNP309 have compromised transcriptional elongation of p53 target genes

Abstract

Abstract MDM2 protein is often found over-expressed in cancers with wild-type p53. A single nucleotide polymorphism (T to G) near the mdm2 P2 promoter, mdm2 SNP309, leads to MDM2 overexpression. This polymorphism is associated with accelerated tumor formation, decreased sensitivity to DNA damage treatment and compromised p53 transcriptional activity. Two G/G SNP309 cell lines MANCA (Burkitt lymphoma) and A875 (melanoma) express wild-type p53 protein not readily degraded by MDM2. This wild-type p53 does not activate target genes even after DNA damage. During the blocked transcription, MDM2 and p53 co-localize at p53 response elements on chromatin. Therefore, we hypothesized that MDM2 localized with p53 on the chromatin was the contributing factor for the compromised p53 transcriptional activity. We explored the mechanism for compromised p53 transcriptional activity in these G/G SNP309 cancer cell lines. When compared to fully activated wild-type p53 in ML-1 cells treated with etoposide, MANCA and A875 cell lines had comparable recruitment of total and initiated RNA pol II at transcription start sites (TSS) for p21 and puma genes. This indicated that G/G SNP309 cells had functional transcription initiation at p53 target genes after DNA damage treatment. We next assessed epigenetic modifications associated with active transcriptional elongation. Using H3K36 trimethylation (H3K36me3) as a hallmark for active elongation, we observed that ML-1 cells had a substantial increase in H3K36me3 after etoposide treatment at p21 and puma TSS that was not seen in MANCA or A875 cell lines. This lack of H3K36me3 suggested an inhibition in early transcriptional elongation. To address the participation of MDM2 in the elongation block we created a stable MDM2 knockdown MANCA cell line. Interestingly, MDM2 knockdown drastically increased MDMX protein levels and moderately increased activation of p53 target genes implying MDMX played a redundant role for MDM2. We are currently investigating the roles of MDM2 and MDMX in blocking transcriptional elongation. Interestingly, we have found that treatment of MANCA cells with low dose Actinomycin D reactivates the p53 pathway and studies are in progress to determine if this is through activation of transcriptional elongation. This work was supported by The Breast Cancer Research Foundation and The National Science Foundation (grant MCB-0744316) to J.B. M.R. was supported by the MBRS-RISE Program to Hunter College 3R25-GM060665. Citation Format: Melissa Rosso, Alla Polotskaia, Jill Bargonetti. Cancer cells with G/G mdm2 SNP309 have compromised transcriptional elongation of p53 target genes. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 468. doi:10.1158/1538-7445.AM2014-468