Abstract 1096: Arf-independent activation of p53 by the Dmp1 tumor suppressor

Abstract

Abstract Cyclin D binding myb-like protein 1 (Dmp1; Dmtf1) is a tumor suppressor that is deleted in ∼40 % of human non-small cell lung carcinomas. The Dmp1 promoter receives signals from oncogenic Ras and the protein shows its activity as a tumor suppressor by directly binding and transactivating the Arf promoter, and thereby inducing Arf-, p53-dependent cell cycle arrest. Both Eµ-Myc and K-rasLA-induced tumor development was accelerated in Dmp1+/− and Dmp1−/− mice, suggesting that Dmp1 is haplo-insufficient for tumor suppression. In Eµ-Myc lymphomas, the combined frequencies of p53 mutation and Arf deletion in the Dmp1+/− or Dmp1−/− mice were significantly lower (∼10 %) than that in Dmp1+/+ littermates (∼50 %), indicating that Dmp1 is a physiological regulator of the Arf-p53 pathway in vivo. We recently found that the frequency of p53 mutation (∼40 %) was significantly decreased in both Dmp1+/− and Dmp1−/− backgrounds (< 10 %) in the K-rasLA lung cancer model where the Ink4a/Arf involvement is very rare. Moreover, our recent data show that Dmp1 physically interacts with p53 to neutralize the activity of Hdm2. To demonstrate the Arf-independent function of Dmp1 on p53 activation in vivo, we injected doxorubicin into wild-type, Arf-null, and Dmp1-null mice, harvested the thymus at 2 hr intervals, and studied the expression of the p53 targets, p21cip1 and bbc3, by real-time PCR and immunohistochemistry. Thymus was chosen as a target tissue since Dmp1 is highly expressed in the medulla in our histochemical studies. Western blotting analyses with DO-1 and phosphoserine-specific antibodies verified p53 activation by doxorubicin. Cleaved caspase 3 was detectable in the thymic medulla of wild-type mice 6 hrs after tail injection of doxorubicin, indicating the induction of apoptotic cell death by the genotoxic drug. Cleaved caspase 3 staining was significantly decreased in the thymus from Arf−/− mice as compared to wild-type mice, but the staining was even lower in Dmp1−/− mice. Consistent with these findings, the p21cip1 mRNA induction (∼23-fold increase in wild-type mice) was more significantly compromised in Dmp1−/− mice (∼7-fold) than in Arf−/− mice (∼15-fold). Bbc was undetectable in Dmp1−/− thymus even 4-6 hrs after injection of doxorubicin while it was significantly induced (∼3-fold) in both wild-type and Arf−/− mice. These mRNA data were confirmed by immunohistochemical staining of thymic tissues with p21cip1 and bbc3-specific antibodies. Together, our data demonstrate the Arf-independent activation of p53 by Dmp1 in response to genotoxic stress in vivo. We are currently conducting the same series of experiments in the mouse lung. 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 1096.