Distribution of topoisomerase II-mediated cleavage sites and relation to structural and functional landmarks in 830 kb of Drosophila DNA.
The pattern of sites for cleavage mediated by topoisomerase II was determined in 830 kb of cloned DNA from the Drosophila X chromosome, with the objectives of comparing it with mapped structural and functional landmarks and examining if the correlations with such landmarks reported in individual loci can be generalized to a region approximately 100 times longer. The relative frequencies of topoisomerase II cleavage sites in 247 restriction fragments from 67 clones were quantified by hybridization with probes prepared from DNA fragments which abutted all cleavage sites in each clone, selected through the covalently bound topoisomerase II subunit; the specificity and quantitative nature of this method were demonstrated using a plasmid DNA model. The 12 restriction fragments with strong nuclear scaffold attachment (SAR) activity, of which seven possess autonomous replication (ARS) activity, show statistically strong coincidence or contiguity ( P </=0.11) with regions of high topoisomerase II cleavage site frequency. These regions show no correlation with repetitive sequence or A/T or C/G content and some extend over >10 kb; their sensitivity is therefore unlikely to be due to alternating purine-pyrimidine repeats or regions of Z conformation, which are preferred motifs. The hypothesis that they possess intrinsic curvature is consistent with the similarity of their length and spacing to regions of predicted curvature in the 315 kb DNA of Saccharomyces cerevisiae chromosome III and with the reported strong binding preference of topoisomerase II for curved DNA. The topoisomerase II cleavage pattern in this DNA further shows that its relationships to functional properties seen in individual loci, especially to MAR/SAR and ARS activity and to the restricted accessibility of DNA to topoisomerase II in vivo, can be generalized to much longer regions of the genome.
Miassod
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Laboratoire de Génétique et Physiologie du Développement, Centre National de la Recherche Scientifique, Case 907, 13288 Marseille, France.
Razin
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Hancock
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D000818:Animals; D001665:Binding Sites; D002874:Chromosome Mapping; D004247:DNA / Q000737:chemistry / Q000378:metabolism*; D004250:DNA Topoisomerases, Type II / Q000378:metabolism*; D004330:Drosophila; D004591:Electrophoresis, Polyacrylamide Gel; D009690:Nucleic Acid Conformation; D015183:Restriction Mapping; D013329:Structure-Activity Relationship
D004247:DNA; D004250:DNA Topoisomerases, Type II