Arsenic Trioxide Augments Chk2/p53-mediated Apoptosis by Inhibiting Oncogenic Wip1 Phosphatase

Junichi Tsukada,Nobuyuki Onishi,Yasuhide Watanabe,Yusuke Tsukuda,Kyoko Toyoshima,Takeshi Kondo,Yuki Hazaka,Akinori Yoda,Yasuhiro Minami,Yoshiya Tanaka

doi:10.1074/jbc.m800560200

Abstract

The oncogenic Wip1 phosphatase (PPM1D) is induced upon DNA damage in a p53-dependent manner and is required for inactivation or suppression of DNA damage-induced cell cycle checkpoint arrest and of apoptosis by dephosphorylating and inactivating phosphorylated Chk2, Chk1, and ATM kinases. It has been reported that arsenic trioxide (ATO), a potent cancer chemotherapeutic agent, in particular for acute promyelocytic leukemia, activates the Chk2/p53 pathway, leading to apoptosis. ATO is also known to activate the p38 MAPK/p53 pathway. Here we show that phosphatase activities of purified Wip1 toward phosphorylated Chk2 and p38 in vitro are inhibited by ATO in a dose-dependent manner. Furthermore, DNA damage-induced phosphorylation of Chk2 and p38 in cultured cells is suppressed by ectopic expression of Wip1, and this Wip1-mediated suppression can be restored by the presence of ATO. We also show that treatment of acute promyelocytic leukemia cells with ATO resulted in induction of phosphorylation and activation of Chk2 and p38 MAPK, which are required for ATO-induced apoptosis. Importantly, this ATO-induced activation of Chk2/p53 and p38 MAPK/p53 apoptotic pathways can be enhanced by siRNA-mediated suppression of Wip1 expression, further indicating that ATO inhibits Wip1 phosphatase in vivo. These results exemplify that Wip1 is a direct molecular target of ATO.

Highlights

Arsenic trioxide (As2O3; arsenic trioxide (ATO))3 has been used therapeutically for several thousand years as a part of traditional Chinese
We show that treatment of acute promyelocytic leukemia cells with ATO resulted in induction of phosphorylation and activation of Chk2 and p38 Mitogen-activated protein kinases (MAPK), which are required for ATO-induced apoptosis
All-trans-retinoic acid (ATRA), another drug for therapy of acute promyelocytic leukemia (APL) patients, failed to induce the phosphorylation of Chk2, p38 MAPK, and p53 in NB4 cells (Fig. 1A). These results indicate that ATO, but not APO or ATRA, induces activation of Chk2/p53 and p38 MAPK/p53 pathways

Summary

Introduction

Arsenic trioxide (As2O3; ATO)3 has been used therapeutically for several thousand years as a part of traditional Chinese. ATO-induced activation of Chk2 and p38 MAPK in NB4 cells was inhibited by NAC (10 mM) as assessed by immunoblot analysis with phosphorylation site-specific antibodies (Fig. 2C, compare lanes 2 and 4).

Results

Conclusion