Radiation-induced heat-labile sites that convert into DNA double-strand breaks

The yield of DNA double-strand breaks (DSBs) in SV40 DNA irradiated in aque ous solution was found to increase by more than a factor of two as a result of postirradiation incubation of the DNA at 50 degrees C and pH 8.0 for 24 h, This is in agreement with data from studies performed at 37 degrees C t hat were published previously. Importantly, similar results were also obtai ned from irradiation of mammalian DNA in agarose plugs. These results sugge st that heat-labile sites within locally multiply damaged sites are produce d by radiation and are subsequently transformed into DSBs, Since incubation at 50 degrees C is typically employed for lysis of cells in commonly used pulsed-field gel assays for detection of DSBs in mammalian cells, the possi bility that heat-labile sites are present in irradiated cells was also stud ied. An increase in the apparent number of DSBs as a function of lysis time at 50 degrees C was found with kinetics that was similar to that for irrad iated DNA, although the magnitude of the increase was smaller. This suggest s that heat-labile sites are also formed in the cell. If this is the case, a proportion of DSBs measured by the pulsed-field gel assays may occur duri ng the lysis step and may not be present in the cell as breaks but as heat- labile sites. It is suggested that such sites consist mainly of heat-labile sugar lesions within locally multiply damaged sites. Comparing rejoining o f DSBs measured with short and long lysis procedure indicates that the heat -labile sites are repaired with fast kinetics in comparison with repair of the bulk of DSBs, (C) 2000 by Radiation Research Society.