Uracil-DNA Glycosylase (UNG)-Deficient Mice Reveal a Primary Role of the Enzyme during DNA Replication

Abstract

Gene-targeted knockout mice have been generated lacking the major uracil-DNA glycosylase, UNG. In contrast to ung- mutants of bacteria and yeast, such mice do not exhibit a greatly increased spontaneous mutation frequency. However, there is only slow removal of uracil from misincorporated dUMP in isolated ung-/- nuclei and an elevated steady-state level of uracil in DNA in dividing ung-/- cells. A backup uracil-excising activity in tissue extracts from ung null mice, with properties indistinguishable from the mammalian SMUG1 DNA glycosylase, may account for the repair of premutagenic U:G mispairs resulting from cytosine deamination in vivo. The nuclear UNG protein has apparently evolved a specialized role in mammalian cells counteracting U:A base pairs formed by use of dUTP during DNA synthesis.

Separation of Function between Isotype Switching and Affinity Maturation In Vivo during Acute Immune Responses and Circulating Autoantibodies in UNG-Deficient Mice

“…However, extrapolating this result to in vivo might not be straightforward, as exemplified by the Msh2 2/2 mice (12, 14-17). The more severe than expected Ab diversification phenotype in Msh2 2/2 mice is likely related to MSH2 roles in cell-cycle checkpoints that presumably affect the viability of germinal center B cells in vivo (15,17), which does not seem to be an issue with UNG as previous results (9,22) and our results indicate. Still, the presumably strong CSR deficiency of Ung 2/2 mice in vivo was not obvious from the available data.…”

Mutation frequencies and AID activation state in B-cell lymphomas from Ung-deficient mice

“…In order to determine single-site mutation frequencies in purified B-cells from lymphomas and controls, nontranslated regions of the p53, bcl-6, and c-myc genes ( Figure 2a Lymphoma mutation patterns in Ung-deficient mice H Nilsen et al controls was apparent in the typical Ung À/À tumors ( Figure 2b). This is in good agreement with the previously noted modest 1.3-1.4-fold increase in mutation frequency in a nontranscribed reporter gene in Ung À/À mice (Nilsen et al, 2000). The unusual 8-Ung À/À tumor showed a sixfold and threefold increased mutation frequencies at the bcl-6 and c-myc loc, respectively (Figure 2b).…”

“…Furthermore, spleen cells from such animals have a slightly increased mutation frequency, apparently due to deaminated DNA cytosine residues (Nilsen et al, 2000). These Ung À/À mice show a perturbed somatic hypermutation pattern in expressed antibody V genes, consistent with reduced excision of uracil from DNA, and class switch recombination is also substantially inhibited (Rada et al, 2002).…”

Uracil in DNA and its processing by different DNA glycosylases

“…They also have a weak preference for U:G contexts over U:A contexts, but this preference is sequence dependent and not absolute (Eftedal et al 1993;Slupphaug et al 1995). Compared with other uracil-DNA glycosylases, UNG2 (and UNG1) has a very high turnover number, compatible with the dominant role of UNG2 in post-replicative repair of incorporated uracil (U:A context), where repair has to keep up with the rapid movement of the replication fork (Otterlei et al 1999;Nilsen et al 2000;Akbari et al 2004). UNG2 also appears to have an important role in the repair of U:G mismatches, at least in human cells in vitro (Kavli et al 2002;Akbari et al 2004).…”