1037 POSTER The Prolyl-3-hydroxylases (P3H) and P3H-related Genes CRTAP and SC65 Are Novel Transcriptionally Silenced Genes in Burkitt's Lymphoma

Abstract

Background: UHRF1 (Ubiquitin-like containing PHD Ring Finger), an anti-apoptotic protein essential for cell proliferation, is over-expressed in several types of cancer. UHRF1 participates in a huge macro-molecular complex including DNMT1 (DNA methyltransferase 1), Tip60 (a histone acetyltransferase), HAUSP (a ubiquitin specific protease) and HDAC1 (a histone deacetylase). It has been shown that HAUSP protects, in vitro, UHRF1 from auto-ubiquitination and regulates its stability in vivo. We previously showed that thymoquinone (TQ), an anti-cancer drug, induced UHRF1 down-regulation by a p73and caspase 3dependent process. The goal of the present study was to determine more precisely the pathway involved in the degradation of UHRF1 by TQ. Material and Methods: Jurkat cells (T lymphoblastic leukaemia cells) and human astrocytoma cells (cell line U87) were used as cancer cell models. Western blot experiments were performed to detect UHRF1, HAUSP and p73 in both cell lines. Results: We have observed that TQ induced a dose-dependent downregulation of UHRF1 and HAUSP accompanied with p73 up-regulation. Interestingly, kinetic study revealed the presence of higher bands of UHRF1 expression as assessed by western blotting on both cell lines. Co-immunoprecipitation experiments allowed us to demonstrate, in Jurkat cells, that these bands were due to ubiquitination of UHRF1. These data show that the degradation of UHRF1, challenged by TQ, is due to its ubiquitination through an as yet unknown mechanism but which appears dependent upon HAUSP down-regulation. Conclusion: In conclusion, we propose that UHRF1 ubiquitination is a key event in the TQ-induced apoptosis in cancer cells. This ubiquitination might result from the auto-ubiquitination activity of UHRF1, following HAUSP down-regulation.