Overexpression of the atypical protein kinase C ζ reduces topoisomerase II catalytic activity, cleavable complexes formation, and drug-induced cytotoxicity in monocytic U937 leukemia cells

Abstract

In this study, we evaluated the influence of protein kinase Cζ (PKCζ) on topoisomerase II inhibitor-induced cytotoxicity in monocytic U937 cells. In U937-ζJ and U937-ζB cells, enforced PKCζ expression, conferred by stable transfection of PKCζ cDNA, resulted in total inhibition of VP-16- and mitoxantrone-induced apoptosis and decreased drug-induced cytotoxicity, compared with U937-neo control cells. In PKCζ-overexpressing cells, drug resistance correlated with decreased VP-16-induced DNA strand breaks and DNA protein cross-links measured by alkaline elution. Kinetoplast decatenation assay revealed that PKCζ 1 overexpression resulted in reduced global topoisomerase II activity. Moreover, in PKCζ-overexpressing cells, we found that PKCζ interacted with both α and β isoforms of topoisomerase II, and these two enzymes were constitutively phosphorylated. However, when human recombinant PKCζ (rHPKCζ) was incubated with purified topoisomerase II isoforms, rH-PKCζ interacted with topoisomerase IIβ but not with topoisomerase IIα. PKCζ/topoisomerase IIβ interaction resulted in phosphorylation of this enzyme and in decrease of its catalytic activity. Finally, this report shows for the first time that topoisomerase IIβ is a substrate for PKCζ, and that PKCζ may significantly influence topoisomerase II inhibitor-induced cytotoxicity by altering topoisomerase IIβ activity through its kinase function.