DNA Double-Strand Break Repair

Abstract

V(D)J recombination has been examined in several X-ray-sensitive and double-strand break repair-deficient Chinese hamster cell mutants. Signal joint formation was affected in four mutants (xrs 5, XR-1, V-3, and XR-V9B cells, representing complementation groups 1 through 4, respectively) defective in DNA double-strand break rejoining. Among these four, V-3 and XR-V9B were the most severely affected. Only in V-3 was coding joint formation also affected. Ataxia telangiectasia-like hamster cell mutants (V-E5 and V-G8), which are normal for double-strand break repair but are X ray sensitive, were normal for all aspects of the V(D)J recombination reaction, indicating that X-ray sensitivity is not the common denominator but that the deficiency in double-strand break repair appears to be. The abnormality at the signal joints consisted of an elevated incidence of nucleotide loss from each of the two signal ends. Interestingly, in complementation groups 1 (xrs 5) and 2 (XR-1), signal joint formation was within the normal range under some transfection conditions. This suggests that the affected gene products in these two complementation groups are not catalytic components. Instead, they may be either secondary or stochiometric components involved in the later stages of both the V(D)J recombination reaction and double-strand break repair. The fact that such factors can affect the precision of the signal joint has mechanistic implications for V(D)J recombination.The exons encoding the immunoglobulin and T-cell receptor variable-region domains are assembled during lymphocyte development by V(D)J recombination. This site-specific reaction is directed by a pair of joining signals recognized by components of the recombination activity. The consensus sequence of each signal consists of a palindromic heptamer and an AT-rich nonamer. These two elements are separated by a 12-base spacer at one signal and a 23-base spacer at the other. The recombination site for each signal is at the end of the heptamer on the side distal from the nonamer. During recombination, coding ends and signal ends are generated by recombinase-mediated cleavage at the crossover sites. The two coding ends are joined to form what is termed a coding joint, and the two signal ends are joined to form a signal joint (reviewed in reference 20). Modification can occur at the two coding ends (20,25) in ways that are similar in some respects to modifications of chromosomal ends left after transposon excisions in eukaryotes (4, 21, 36).Murine scid (3) has been shown to be defective in V(D)J recombination, defective in the repair of double-strand breaks, and sensitive to X-irradiation (la, 2, 4a, 10, lOa, 28, 31, 32). The scid defect in double-strand break repair implies that the processing of coding ends in the V(D)J recombination reaction is similar to the processing of chromosomal breaks. Both processes can involve nucleotide loss, and both processes can involve nucleotide addition (22, 27).The complexity of mechanisms involved in cellular responses to ionizing radiati...