Phosphorylation of serine 1106 in the catalytic domain of topoisomerase IIα regulates enzymatic activity and drug sensitivity

Topoisomerases alter DNA topology and are vital for the maintenance of genomic integrity. Topoisomerases I and II are also targets for widely used antitumor agents. We demonstrated previously that in the human leukemia cell line, HL-60, resistance to topoisomerase (topo) II-targeting drugs such as etoposide is associated with site-specific hypophosphorylation of topo IIalpha. This effect can be mimicked in sensitive cells treated with the intracellular Ca2+ chelator, 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA-AM). Here we identify Ser-1106 as a major phosphorylation site in the catalytic domain of topo IIalpha. This site lies within the consensus sequence for the acidotrophic kinases, casein kinase I and casein kinase II. Mutation of serine 1106 to alanine (S1106A) abrogates phosphorylation of phosphopeptides that were found to be hypophosphorylated in resistant HL-60 cells or sensitive cells treated with BAPTA-AM. Purified topo IIalpha containing a S1106A substitution is 4-fold less active than wild type topo IIalpha in decatenating kinetoplast DNA and also exhibits a 2-4-fold decrease in the level of etoposide-stabilized DNA cleavable complex formation. Saccharomyces cerevisiae (JN394t2-4) cells expressing S1106A mutant topo IIalpha protein are more resistant to the cytotoxic effects of etoposide or amsacrine. These results demonstrate that Ca2+-regulated phosphorylation of Ser-1106 in the catalytic domain of topo IIalpha modulates the enzymatic activity of this protein and sensitivity to topo II-targeting drugs.