Abstract
Arsenic Trioxide (As2O3) has major efficacy in the treatment of acute promyelocytic leukemia (APL), but its use in other malignancies is limited by the need for high intracellular concentrations to induce apoptosis. Prior work in our laboratory has demonstrated that the p38 MAP kinase (MAPK) pathway is activated following treatment of cells with As2O3 and exhibits negative regulatory effects on As2O3-induced apoptosis and growth suppression. In the current study, we sought to identify upstream effector mechanisms by which the p38 pathway is activated by As2O3 in leukemic cells. We found that the MAPK kinase kinase TAO2 (thousand and one amino acid protein kinase 2) is phosphorylated on Ser181 after treatment of NB4, NB4.306, and U937 cells with arsenic. Such phosphorylation was rapid, occurring as early as after 5 minutes of As2O3 treatment. In addition, our data indicate that such phosphorylation occurs downstream of As2O3-induced redox reactions, as demonstrated by increased phosphorylation in cells pretreated with the oxidizing agent buthionine sulfoximine (BSO) and decreased phosphorylation following pretreatment with the reducing agent dithiothreitol (DTT). Arsenic treatment of the cells also resulted in activation of the kinase domain of TAO2, as evidenced in in vitro kinase assay studies using ATF2 as an exogenous substrate. siRNA-mediated TAO2 knockdown resulted in inhibition of As2O3-induced p38 phosphorylation, suggesting that this kinase acts as an upstream effector of the arsenic-activated p38 MAPK pathway. Moreover, in studies to determine the functional relevance of TAO2 in the induction of As2O3-dependent antileukemic responses we found that siRNA-mediated TAO2 knockdown enhanced the suppressive effects of As2O3 on KT1-derived leukemic progenitor (CFU-L) growth in clonogenic assays in methylcellulose. Altogether, our data demonstrate that TAO2 is activated during arsenic treatment of leukemic cells lines and acts as an upstream activator of the p38 MAPK pathway. Such activation appears to occur in a negative feedback regulatory manner to compensate for the suppressive effects of As2O3 on leukemic cell growth. Importantly, these findings raise the possibility that targeting TAO2 may provide a novel approach to enhance the generation of the antileukemic properties of As2O3.
Glibenclamide–sulfonylurea receptor 1 antagonist alleviates LPS-induced BV2 cell activation through the p38/MAPK pathway