TOPOISOMERASE-I ACTION ON THE HETEROCHROMATIC DNA FROM THE BRINE SHRIMP ARTEMIA-FRANCISCANA - STUDIES IN-VIVO AND IN-VITRO

The genomes of higher eukaryotes contain various amounts of tandem rep eated DNA sequences (satellite DNA) typically located in the constitut ive heterochromatin, the most highly condensed region of interphase ch romosomes. We have previously demonstrated that an AluI DNA family of repeats is the major component of constitutive heterochromatin in the brine shrimp Artemia franciscana. The analysis of cloned heterochromat ic fragments revealed that this repetitive DNA shows a stable curvatur e conferring a solenoidal geometry to the double helix. In this paper we provide evidence, using the antitumour drug camptothecin, that, in vivo, topoisomerase I cleaves heterochromatin with a frequency compara ble with that observed in the whole genome. The analysis of the break sites shows that the enzyme cleaves heterochromatic DNA at specific si tes characterized by a degenerate consensus sequence. Moreover the enz yme-mediated breaks have, in vitro, a degenerate consensus sequence si milar to, but not identical with, the in vivo one. Some of these sites are influenced by the DNA flanking the heterochromatic insert, sugges ting that structural variations could modify the enzyme specificity.