Elsevier

DNA Repair

Volume 6, Issue 4, 1 April 2007, Pages 505-516
DNA Repair

Uracil in DNA—General mutagen, but normal intermediate in acquired immunity

https://doi.org/10.1016/j.dnarep.2006.10.014Get rights and content

Abstract

Deamination of cytosine in DNA results in mutagenic U:G mispairs, whereas incorporation of dUMP leads to U:A pairs that may be genotoxic directly or indirectly. In both cases, uracil is mainly removed by a uracil-DNA glycosylase (UDG) that initiates the base excision repair pathway. The major UDGs are mitochondrial UNG1 and nuclear UNG2 encoded by the UNG-gene, and nuclear SMUG1. TDG and MBD4 remove uracil from special sequence contexts, but their roles remain poorly understood. UNG2 is cell cycle regulated and has a major role in post-replicative removal of incorporated uracils. UNG2 and SMUG1 are both important for prevention of mutations caused by cytosine deamination, and their functions are non-redundant. In addition, SMUG1 has a major role in removal of hydroxymethyl uracil from oxidized thymines. Furthermore, UNG-proteins and SMUG1 may have important functions in removal of oxidized cytosines, e.g. isodialuric acid, alloxan and 5-hydroxyuracil after exposure to ionizing radiation. UNG2 is also essential in the acquired immune response, including somatic hypermutation (SHM) required for antibody affinity maturation and class switch recombination (CSR) mediating new effector functions, e.g. from IgM to IgG. Upon antigen exposure B-lymphocytes express activation induced cytosine deaminase that generates U:G mispairs at the Ig locus. These result in GC to AT transition mutations upon DNA replication and apparently other mutations as well. Some of these may result from the generation of abasic sites and translesion bypass synthesis across such sites. SMUG1 can not complement UNG2 deficiency, probably because it works very inefficiently on single-stranded DNA and is down-regulated in B cells. In humans, UNG-deficiency results in the hyper IgM syndrome characterized by recurrent infections, lymphoid hyperplasia, extremely low IgG, IgA and IgE and elevated IgM. Ung−/− mice have a similar phenotype, but in addition display dysregulated cytokine production and develop B cell lymphomas late in life.

Introduction

The RNA base uracil may be present in DNA in small amounts as a result of cytosine deamination or misincorporation of dUMP during DNA replication. Such lesions are normally faithfully repaired by base excision repair (BER) initiated by a uracil-DNA glycosylase [1]. Contrary to expectations, mice deficient in the major uracil-DNA glycosylase encoded by the Ung-gene are viable, fertile and develop normally [2]. This is probably due to the existence of at least three alternative uracil-DNA glycosylases named SMUG1 (single strand selective monofunctional uracil-DNA glycosylase), TDG (thymine/uracil mismatch DNA glycosylase) and MBD4 (methyl binding domain 4 protein) [1]. These apparently have specialized functions but may also serve as backups for each other. Surprisingly, uracil in DNA and uracil-DNA glycosylase encoded by the UNG gene also have functions in the acquired immune system [3], [4], as well as in the innate immune system in defense against retroviral infections [5]. Thus, there is important crosstalk between the ancient DNA repair mechanisms and the innate immune system, as well as the much younger acquired immune system that came with vertebrates. For the acquired immune system mismatch repair proteins and proteins required for double strand break repair are also involved. This demonstrates that the interactions between these defense systems are in fact quite extensive [6]. While Ung-deficient mice have no overt phenotype macroscopically, they develop B-cell lymphomas late in life [7], [8]. In this review we will present the current knowledge about the origins of uracil in DNA and discuss functional characteristics of the quantitatively dominating glycosylases removing uracil and some closely related oxidative lesions. Finally, we will discuss the role of uracil and uracil-DNA glycosylase UNG2 in the Ig diversification process.

Section snippets

Uracil in DNA

Uracil in DNA is in several ways a rather special case among DNA lesions. First, it is a normal constituent in RNA but not in cellular DNA. Second, template uracil in U:G mispairs is a non-blocking, but 100% mutagenic lesion that results in mutation in one of the two daughter cells if it is replicated across. Incorporation of dUMP from the normal nucleotide dUTP results in U:A pairs that themselves are not mutagenic. However, chromosomal abasic sites resulting from uracil-removal are mutagenic

Removal of uracil by uracil-DNA glycosylases

At least four different uracil-DNA glycosylases (UDGs) have been identified in mammalian cells. These are UNG1/UNG2, SMUG1, TDG and MBD4 [30]. In addition, mouse NEIL1 has also been demonstrated to excise uracil and thymine from mismatches [31], but the biological significance of this remains unclear. Among the UDGs, UNG, SMUG1 and TDG belong to the same protein superfamily, possess the same fold and have probably evolved from the same ancestral gene [32]. Although our knowledge of the

Role of uracil and UNG in immunoglobulin gene diversification

The acquired immune system performs its main function through immunoglobulin (Ig) molecules. A primary repertoire of low affinity antibodies (IgM) is generated by the RAG1/RAG2 dependent V(D)J recombination process in the bone marrow [62]. The Ig based defense mechanism is adaptive and responds dynamically during the course of infection. This is achieved by three additional somatic Ig gene diversification processes; gene conversion (GC), somatic hypermutation (SHM) and class switch

Future perspectives

Although our understanding of the uracil-DNA glycosylases has advanced considerably over the last decade, several aspects remain to be elucidated. A major question is how UNG2 may initiate either repair of uracil, or induce mutations or recombination via SHM and CSR, respectively. It is very likely that UNG2, as well as many of the other factors involved in these processes are subject to extensive post-translational regulation that direct assembly of specific and higher order protein complexes.

Acknowledgements

This work was supported by The Research Council of Norway, the National Program in Functional Genomics (FUGE) administered by The Research Council of Norway, The Norwegian Cancer Society, The Cancer Fund at St.Olav's Hospital, Trondheim, The Svanhild and Arne Must Fund for Medical Research and The European Union Program DNA Repair.

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