ChIP-on-chip analysis of DNA topoisomerases.

Here we describe an adapted ChIP-on-chip protocol for the analysis of DNA topoisomerase chromosomal binding in Saccharomyces cerevisiae cells. The ChIP-on-chip technique is based on the immunoprecipitation of crosslinked chromatin (ChIP, chromatin immunoprecipitation), followed by DNA amplification and hybridization to high-density oligonucleotide arrays (Chip). Comparison of the signal intensities of immunoprecipitated and control fractions provides a measurement of the protein-DNA association along entire genomes.

Top1- and Top2-mediated topological transitions at replication forks ensure fork progression and stability and prevent DNA damage checkpoint activation.

DNA topoisomerases solve topological problems during chromosome metabolism. We investigated where and when Top1 and Top2 are recruited on replicating chromosomes and how their inactivation affects fork integrity and DNA damage checkpoint activation. We show that, in the context of replicating chromatin, Top1 and Top2 act within a 600-base-pair (bp) region spanning the moving forks.