NUCLEAR-STRUCTURE AND THE MICRODISTRIBUTION OF RADIATION-DAMAGE IN DNA

Evidence for the roles of proteins and metal ions in the microheteroge neity of DNA damage is reviewed. Decondensation of chromatin in hypoto nic buffers markedly sensitizes the DNA to radiation, while treatment of nuclei with hypertonic buffers strips the DNA of histones and other nuclear proteins and enhances the radiosensitivity of the DNA with re spect to double-strand break (dsb) formation. Addition of the radical scavenger DMSO reduces the yield of strand breaks, but dehistonized ch romatin remains similar to 2.5 times more sensitive to radiation than does native chromatin at 0.1 M DMSO. DNA-protein crosslink (DPC) forma tion is relatively unaffected by the removal of the majority of histon es from chromatin. Most DPC form at or near the nuclear matrix, and ma trix is stabilized and radiosensitized by Cu++. To elucidate the role of Cu++, the induction of dsb and DPC by gamma-radiation has been comp ared with that by hydroxyl radical from Fe++-EDTA, or Cu++ catalysed F enton reactions. Data comparing the size of DNA fragments produced, th e effect of expanding or dehistonizing chromatin, and the effects of r adical scavengers suggest that gamma-radiation and Fe++-EDTA produce d sb at open chromatin sites, whereas Cu++-generated dsb are similar to radiation-induced DPC in their location at the nuclear matrix. Both me tal ions appeared to produce damage by site-specific generation of hyd roxyl radicals. The nuclear matrix, the proteinaceous skeleton which a nchors chromosomal loops and provides sites for DNA replication and tr anscription, binds metal ions and matrix-attachment DNA regions (MARs) consisting of 300+ bp of AT-rich DNA. The interaction of cloned MARs with isolated nuclear matrices has been found to be hypersensitive to crosslinking upon gamma-irradiation, in comparison with associations f ormed by similarly sized DNA fragments lacking MAR sequences. Thus, th e non-random distribution of radiation damage is partially explained b y the protection of DNA afforded by histones and chromatin structure a nd partially by the hypersensitivity of DNA-nuclear matrix association s.