DRUG-SENSITIVITY AND DNA-BINDING OF A SUBFORM OF TOPOISOMERASE II-ALPHA IN RESISTANT HUMAN HL-60 CELLS

Topoisomerase II alpha (170 kDa) expressed in human HL-60 cells is het erogeneous in charge. By two-dimensional electrophoresis and chromatof ocussing two major subforms with pi of 6.5 and 6.7 can be resolved. By preparative anion-exchange chromatography we separated the known topo isomerase II isoenzymes (170/180 kDa) and in addition a late-eluting 1 70 kDa form, which has not been described before. The catalytic optimu m of this late-eluting form is shifted to pH 9.4. It is more than 100- fold resistant to orthovanadate, amsacrine or etoposide, and has an in creased salt stability. SDS-treatment induces covalent attachment of t his enzyme fraction to calf thymus DNA in the absence of drug. The lat ter observations indicate an increase in DNA-binding. In the tightly D NA-bound state the late-eluting enzyme is not targeted by cleavable co mplex forming drugs. Accordingly, cells may become drug-resistant by e xpressing this form predominantly.