Topoisomerase II beta in the cellular response to DNA damage

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Our DNA, with ~3 billion base pairs, encodes the ‘blueprint’ for all the structures and functions of a cell over its lifespan. Maintaining the integrity and stability of this DNA is of paramount importance to survival of the cell and the entire organism. With each cell division, this DNA must be faithfully copied and equally distributed into daughter cells. It must also be prepared to react to any DNA damaging events. Failure to do so can lead to the accumulation of mutations in our DNA, cell aging or cell death. Astoundingly, each cell in a living organism endures tens of thousands of DNA insults every day. These arise spontaneously from both endogenous (from within the body) and exogenous (from outside the body) sources. Paradoxically, oxygen, although absolutely essential for life, is converted to reactive oxygen species as by-products of cell metabolism; these can alter DNA in numerous ways, potential leading to mutations or broken DNA fragments. Naturally occurring radiation found in the earth’s crust and exposure from space, airline travel and medical x-rays can lead to breaks in our DNA. Thankfully, over 200 mammalian genes are known to be involved in the detection and repair of DNA damage. Perhaps paradoxically, other proteins critical for cell function can react to DNA damage and become trapped on the DNA. This is a potential hazard, as these proteins could add to the damage burden, or slow the repair of damage sites. Characterization of these protein behaviours is an area of intense investigation. This application contributes to the field by investigating this behaviour for one enzyme, topoisomerase IIß (TOP2ß). The topoisomerases are an evolutionarily conserved family of enzymes that are involved in untangling and unwinding DNA to allow for its replication (copying) and segregation (separation) into daughter cells. They are found in all organisms, including bacteria, viruses, plants and animals. Although topoisomerases (including the TOP2ß form) have been extensively studied, emerging data from my laboratory points to an as yet uncharacterized role for this protein in the response to damage caused by radiation. In our laboratory, we have developed experimental tools that position us to determine the role TOP2ß plays in the cell’s response to radiation. Using these unique reagents, together with additional resources obtained through international collaborations, we will specifically investigate how TOP2ß becomes trapped on DNA. Using engineered mutant forms of TOP2ß, we will clarify the requirement for enzyme activity and study the kinetics of its movement. Deciphering how our cells respond to the DNA damage, and identification and characterization of new proteins that respond to this damage, is of importance in understanding DNA repair in a wide range of organisms and for identifying targets for the development of agents that can help preserve the long-term stability of our DNA.