strand displacement and subsequent DNA synthesis, which might otherwise result in local expansion of sequence repeats (Tishkoff and others 1997; Freudenreich and others 1998). The temporary inefficiency of this process during early mammalian development could explain the origin of several human syndromes that are associated with expansion of triplet repeats in relevant genes.
A series of pairwise interactions between the relevant proteins in BER seem to occur in most cases without any direct strong protein-protein interactions in the absence of DNA. The XRCC1-LIG3 heterodimer is the only preformed complex, and no large preassembled multiprotein BER complex is likely to exist. Nevertheless, the consecutive ordered interactions may protect reaction intermediates and ensure efficient completion of the correction process after initial DNA damage recognition.
The great majority of endogenous DNA lesions produced by reactive oxygen species are corrected by the BER pathway, and the contributions of the different pathways of nucleotide-excision repair (NER) and mismatch repair are very minor. However, exposure of DNA or cells to ionizing radiation under hypoxic conditions causes the formation of 5′, 8-purine cyclodeoxynucleosides. This chemically stable and distorting form of DNA damage, in which the purine is attached by two covalent bonds to the sugar-phosphate backbone, can be removed only by NER (Heyer and others 2000; Kuraoka and others 2000). Similarly, a major lipid peroxidation product, malondialdehyde, reacts with G to produce an exocyclic pyrimidopurinone (M1G) that requires NER for repair. These are not the major mutagenic or cytotoxic lesions that occur as a consequence of exposure to ionizing radiation, but they could be critical in individuals with impaired ability to perform NER.
Reactive oxygen species cause DNA strand breaks by destroying deoxyribose residues. Such SSBs are processed and repaired by the same enzymes responsible for the later stages of BER, sometimes with the additional steps of exonucleolytic removal of base pairs and phosphorylation of 5′ termini by DNA kinase. In contrast to the continuous protection of DNA reaction intermediates when an altered base residue is replaced however, the initial strand break is fragile and attracts unwelcome recombination events. An abundant nuclear protein, poly(ADP-ribose) polymerase-1 (PARP1), appears to have as its main role the temporary protection of DNA single-strand interruptions (Le Rhun and others 1998; Lindahl and Wood 1999). PARP1 rapidly shuttles strand breaks in DNA on and off, with NAD-dependent synthesis of poly(ADP-ribose) as its release mechanism. PARP1 knockout mice are viable but show increased numbers of spontaneous sister-chromatid exchanges and sensitivity to ionizing radiation. Extracts of cells from such mice contain low concentrations of other PARP enzymes, which may have distinct unknown roles but could also have backup functions. Crossing PARP1 knockout mice with severe combined immunodeficient disease knockout mice that lack DNA-dependent protein kinase, which is required for VDJ recombination during lymphocyte development, alleviates the DNA-processing defect in the latter and allows some low-fidelity recombination (Morrison and others 1997). PARP1 plays no clear role in the BER process itself, as POL β and LIG3 do, but it interacts with the scaffold protein XRCC1 and may in this way accelerate the recruitment of these repair enzymes for strand interruptions (Mackey and others 1999).
Exposure of DNA to ionizing radiation produces about 5–7% as many DSBs as SSBs (e.g., see earlier discussion of Nikjoo and others 1997, 2000). DSBs are sites at which a surprisingly large number of proteins can bind, carry out strand-break repair, and initiate a complex series of cellular signals that regulate cell cycle progression and the induction and activation of many downstream genes. Cells often encounter DNA DSBs under natural circumstances. These include termini (e.g., telomeres at chromosome ends); recombination intermediates; and immunoglobulin rearrangement during the processing of antibody genes (which leads to increased versatility in the repertoire of immature immunocytes), during the processing of stalled or collapsed replication forks arrested by damage on the template strand and during topoisomerase action on DNA. DSB repair enzymes have been suggested as playing an essential role in telomere maintenance in normal undamaged cells (Blackburn 2000).
One critical difference between metabolically generated DSBs and those generated by ionizing radiation is that some fraction of the latter contain complex radiochemical damage that results in LMDS. LMDS (clustered damage) involve frank breaks, radiolytic fragments as termini, and base damage that is processed into breaks by cellular glycosylases (Blaisdell and Wallace 2001). DSBs thus are not inherently novel, although substantial differences between natural and radiation-induced breaks are likely. Cells contain many genes that code for DNA-binding proteins and signal transduction pathways that respond specifically to DNA double-strand breakage. Consequently, cells can distinguish between a naturally occurring end of DNA at a telomere or recombination structure, for example, and a DSB at an unusual location with atypical chemistry. This suggests that metabolic responses to DSBs and LMDS are highly evolved in most cell types and that cells are not completely unprepared and unequipped for these kinds of lesions, but are in fact able to exercise considerable discrimination in their detection and repair. Cells can also repair damage by novel chemicals, such