Investigation of the chromatin remodelling enzyme Uls1 and its interactions with Topoisomerase 2 in S. cerevisiae

Abstract

Acriflavine (ACF) is a Topoisomerase 2 (Top2) poison, a class of drugs which stall Top2 during its reaction cycle causing the formation of persistent DNA breaks to which Top2 remains covalently bound. Deletion of ULS1 causes sensitivity to ACF, with cells showing activation of the Rad53 DNA damage checkpoint. Uls1 is a chromatin remodelling enzyme also implicated in the regulation of levels of SUMO conjugated proteins. We show that Uls1 has both a genetic and physical interaction with Top2, with uls1Δ sensitivity to ACF being linked to Top2 activity. Analysis of Uls1 and Top2 localisation genome wide via ChIP-seq reveals areas where the two proteins co-localise, with Top2 enrichment on chromatin being altered upon deletion of ULS1. At these areas, the presence of Uls1 prevents accumulation of Top2 upon addition of ACF. Our data suggests that Uls1 is required for regulation of stalled Top2. Top2 poisons are used therapeutically as anti-cancer drugs, however these drugs have been implicated in the formation of secondary cancers due to chromosomal translocations arising during the repair of Top2 generated double strand breaks (DSB). The use of dual targeted therapies where a Top2 poison is paired with an inhibitor of another pathway that increases sensitivity to the Top2 poison allows a lower dose to be used, therefore reducing harmful side effects. Our work looked to identify Top2 poison sensitive pathways in S. cerevisiae, where non-essential and essential gene mutants were assayed for sensitivity to ACF. This allowed a comprehensive analysis of 83% of the genes in S. cerevisiae, identifying novel genes within the areas of DNA repair, DNA replication, transcription, chromatin structure, protein modification/degradation, cell division/cell cycle and cellular organisation/cytoskeleton as being important in the response to this bulky adduct.