Abstract 1088: Differences in p53 signaling following induction of nucleotide pool deficiencies with hydroxyurea or PALA

Abstract

Abstract The tumor suppressor p53 is stabilized following the induction of deficiencies in nucleotide pool levels by antimetabolites. N-(phosphonacetyl)-L-aspartate (PALA) reversibly inhibits aspartate transcarbamylase, causing interruption of de novo pyrimidine synthesis; hydroxyurea (HU) reversibly inhibits ribonucleotide reductase, inhibiting the conversion of ribonucleotide triphosphates to deoxynucleotide triphosphates. When HCT116, a human colon carcinoma cell line, is treated with either HU or PALA, p53 accumulates and cells arrest in S-phase. Beyond these similarities, the cellular responses to PALA and HU are strikingly different. Chk1 is phosphorylated to a far greater extent in HU-treated HCT116 cells than following PALA treatment. The lack of Chk1 phosphorylation observed during PALA treatment was found to be due to the p53-dependent transcriptional activation of the Wip1 phosphatase, rather than the inactivity of ATR. Chromatin immunoprecipitation studies performed on PALA-treated cells demonstrated the presence of p53 at the Wip1 promoter, and both Wip1 transcript and protein levels were elevated. In contrast, the residency of p53 over the Wip1 promoter was not observed during HU treatment, nor was Wip1 protein or transcript elevated. During PALA treatment of the isogenic p53 null HCT116 cell line, Wip1 transcript levels are not elevated and Chk1 phosphorylation is restored, demonstrating the role of p53 in the elevation of Wip1 following PALA treatment. Although p53 accumulates during both HU and PALA treatment, the responses to the individual drugs differ so greatly that PALA-treated cells undergo a p53-dependent apoptosis while HU-treated cells enter a cytostatic cell cycle arrest. The post-translational modifications of p53 are dissimilar in the two treatments suggesting a variation of signaling to p53 as well as the downstream actions of the modified p53. The accumulated p53 in PALA-treated cells is phosphorylated at serine 46, an event needed to direct p53 to pro-apoptotic promoters; this phenomenon is not observed in HU-treated cells. Hence, during de novo pyrimidine synthesis inhibition, p53 promotes Wip1 transcription causing dephosphorylation of Chk1 leading to apoptosis. When deoxynucleotide triphosphate synthesis is inhibited, Chk1 is phosphorylated promoting cellular arrest in S-phase. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 1088.