PROJECT SUMMARY DNA topoisomerases are types of enzymes that can specifically resolve topological stresses by transiently introducing strand breaks into DNA molecules and enabling the rotation of the supercoiled DNA strand. Mammalian cells encode two types of topoisomerases: type I topoisomerases (TOP1, TOP1mt, TOP3A, and TOP3B), which introduce single strand breaks into DNA, and type II topoisomerases (TOP2A, TOP2B, and SPO11), which introduce double strand breaks (DSBs) into DNA. This proposal focuses on type II topoisomerases, i.e. TOP2A/2B, in human somatic cells. During cleavage reaction, the tyrosine in the catalytic active site of TOP2 is covalently linked to the DNA backbone and forms the so-called topoisomerase II cleavage complex (TOP2cc). Under normal conditions, TOP2cc forms transiently and is not detectable. However, a wide variety of topoisomerase poisons, including etoposide, have been developed and used as chemotherapeutic drugs for cancer treatment. Mechanistically, etoposide acts to stabilize TOP2cc, which eventually lead to DNA strand breaks and kill tumor cells. While many investigators including us investigated TOP2-induced DNA lesions and how they can be repaired by different repair pathways, this proposal focuses on a new concept that cells have evolved distinct pathways to avoid and limit DNA lesions induced by TOP2. In this proposal, we will determine mechanistically how several unique TOP2 regulators act together to avoid DNA damage and therefore promote cell survival. Results from these studies are critically important for the understanding of therapeutic response to etoposide and other anti-cancer agents. .