The repair of DNA double-strand breaks in mammalian cells and the organization of the DNA in their chromosomes.

The molecular weight of native DNA has been accurately determined by the use of a semiautomated sucrose gradient system. A mondisperse size distribution (speed dependence free) of eighth-of-a-chromatid pieces [1.7 S 10(10) daltons, with 95% confidence (fiducial) limits of +/- 48%] was found. This size has been confirmed by viscoelastometry. Ionizing radiation rapidly breaks each of these pieces into about 21 subunits (again monodisperse) of 8 X 10(8) daltons each. With increaseing dose (greater than 2 krad) the subunits are themselves randomly broken down into even smaller pieces. Postirradiation incubation at 37 degrees C permits the cells to repair both DNA double-strand breaks and intersubunit linkages at the same dose-independent rate (T37) of about 55 min, the same rate as found in Micrococcus radiodurans. The repair data are compatible with a first-order-kinetics repair system, analogous to the post-UV excision-repair system, which becomes saturated at high doses (greater than 60 krad). Specially constructed "enzyme" gradients show that the linkages contain at least two protein molecules each covalently bound to the end of a subunit and linking the subunits together by a disulfide bond(s). Correlation of cell survival and DNA break kinetics yields two possible models. These are that the two-thirds of the lethal events which are due to improperly or unrepaired double-strand breaks result from either (1) a misrepair frequency of 3.5 X 10(-3) (rather high for a mutation frequency) or (2) the induction of a double-strand break in a single eighth-of-a-chromatid unit which is essential for survival but which cannot be repaired, possibly because the unit contains the double-strand break repair system gene(s).