SYNCHRONOUS BLOCK IN DNA-SYNTHESIS INITIATION WITH CHANGE IN CHROMATIN TOPOLOGY MEDIATED BY VP16

The impact of chromatin topology on the DNA synthetic process was stud ied in the human squamous-cell carcinoma cell line SQ-20B, A 1-h expos ure less than or equal to 10 mu M VP16 produced an increase in DNA sup ercoil tension, measured by recording laser light scatter from salt-ex tracted nuclei, This change was precisely paralleled by a decrease in DNA synthesis. The effects on both DNA supercoiling and DNA synthesis were suppressed at VP16 concentrations between 10 and 20 mu M. The cha nges in DNA supercoiling and synthesis at VP16 concentrations less tha n or equal to 10 mu M were eliminated by coincubation with mimosine, a DNA synthesis initiator poison. We conclude that brief exposure to lo w concentrations of VP16 disturbs the balance of torsional energy with in discrete replicon domains by affecting normal topoisomerase II acti vity at sites of replication initiation. The resultant increase in neg ative supercoil tension mediates a topologic checkpoint, limiting the initiation of DNA synthesis. Such a checkpoint may be a common pathway for control, both during the normal replicative cycle and subsequent to DNA damage.