ATM phosphorylates ZBP-89 at Ser202 to potentiate p21waf1 induction by butyrate

Histone deacetylase inhibitors (HDACi) induce growth arrest and differentiation, particularly in the colon where they are potential chemotherapeutic agents. A key mediator of HDACi action is the cyclin dependent kinase (CDK) inhibitor p21(waf1). HDACi treatment of colonic cells promotes the formation of an ATM/ZBP-89/p300 complex on p21(waf1) proximal promoter, and this multi-molecular complex plays an important role in HDACi induction of p21(waf1) expression in vitro and mucosal protection in vivo. Here we found that ZBP-89 is phosphorylated by ATM kinase in vitro and in vivo. Disruption of the ATM phosphorylation motif (202)SQ within the zinc finger domain of ZBP-89 attenuated its ability to enhance p21(waf1) activation by butyrate. Moreover, disruption of the ATM phosphorylation site abrogated the ability of ZBP-89 to potentiate butyrate induction of endogenous p21(waf1) expression. These results demonstrate that ATM phosphorylation of ZBP-89 contributes to HDACi induction of p21(waf1) gene expression.     

Interaction of p21(CDKN1A) with PCNA regulates the histone acetyltransferase activity of p300 in nucleotide excision repair

The cell-cycle inhibitor p21^CDKN1A has been suggested to directly participate in DNA repair, thanks to the interaction with PCNA. Yet, its role has remained unclear. Among proteins interacting with both p21 and PCNA, the histone acetyltransferase (HAT) p300 has been shown to participate in DNA repair. Here we report evidence indicating that p21 protein localizes and interacts with both p300 and PCNA at UV-induced DNA damage sites. The interaction between p300 and PCNA is regulated in vivo by p21. Indeed, loss of p21, or its inability to bind PCNA, results in a prolonged binding to chromatin and an increased association of p300 with PCNA, in UV-irradiated cells. Concomitantly, HAT activity of p300 is reduced after DNA damage. In vitro experiments show that inhibition of p300 HAT activity induced by PCNA is relieved by p21, which disrupts the association between recombinant p300 and PCNA. These results indicate that p21 is required during DNA repair to regulate p300 HAT activity by disrupting its interaction with PCNA.     

p21waf1/Cip1 partially mediates apoptosis in hepatocellular carcinoma cells

p21waf1/Cip1 (p21) is a tumor suppressor gene involved in apoptosis in many cancer cell types induced by different agents. In spite of concomitant induction of p21 by many anti-cancer drugs, including inhibitors of histone deacetylases (HDACi), its pro-apoptotic action has been debated during the last several years due to a lack of direct evidence regarding the exact role of p21 in apoptosis. With the help of anti-sense p21 expression, we show here that p21 is mediating the apoptotic effects of HDACi 4-phenylbutyrate (4-PB) and Trichostatin A (TSA) on the hepatocellular hepatocarcinoma Hep3B cells. Hep3B cells were transfected by EGFP-p21 anti-sense or sense plasmids, and apoptosis induced by HDACi was assessed by TUNEL assay. The results show that the p21 anti-sense construct prevents apoptosis, induced by HDAC inhibitors in Hep3B cells. The obtained results suggest an important role for p21 in mediating the apoptotic effect of HDACi.