Nucleotide Excision Repair

https://doi.org/10.1016/S0079-6603(04)79004-2Get rights and content

Publisher Summary

DNA damage is a common occurrence that compromises the functional integrity of DNA. It is not surprising then that cells have multiple mechanisms for coping with DNA damage, including those introduced by sunlight and other environmental agents. Nucleotide excision repair is a pluripotent pathway for the recognition and removal of a broad spectrum of DNA lesions. Nucleotide excision repair involves the removal of damaged DNA bases by dual incisions bracketing the damaged base, release of the damaged base in the form of 12–13 nucleotide-long oligomers in prokaryotes and 24– 32 nucleotide-long oligomers in eukaryotes followed by polymerase-mediated replacement of the excised nucleotides and sealing the repair patch with ligase. Structural work currently being carried out by numerous groups is expected to provide an understanding at the atomic level for both prokaryotic and eukaryotic excision nucleases, which will aid in designing further biochemical experiments to refine current models.

Introduction

DNA damage is an important event in the initiation and progression of cancer. DNA is a highly reactive macromolecule that is a target for both physical and chemical agents that alter the base(s) and/or the phosphate backbone. Lesions in DNA at the time of replication may be mutagenic and can lead to cancer or cell death. All organisms have elaborate cellular responses to DNA-damaging agents, including both tolerance and repair mechanisms. A critical component of the cellular response to DNA damage includes the repair pathways dedicated to correcting damage or errors in DNA. Nucleotide excision repair (excision repair)1 is a versatile pathway that recognizes and removes a wide spectrum of DNA lesions. In eukaryotes, components of excision repair are also involved in other repair pathways and in various aspects of DNA metabolism.

This chapter gives a general discussion of cellular responses to DNA-damaging agents before presenting a detailed discussion of nucleotide excision repair in Escherichia coli and humans as examples of this repair pathway in prokaryotes and eukaryotes. The basic strategy of this essential repair pathway is conserved from E. coli to humans, but the proteins are not conserved and there are some differences in the mechanistic details. DNA repair is modulated by both transcription and condensation into chromatin. Transcription stimulates excision repair both in E. coli and in humans in a process dependent on proteins called transcription-repair coupling factors. The structural organization of chromosomes into densely compacted nucleosomes has a profound effect on DNA repair. This inhibitory effect of chromatin on repair is alleviated by chromatin remodeling and accessibility factors.

Excision repair genes have been found in all free-living organisms. A defect in nucleotide excision repair causes extreme ultraviolet (UV) sensitivity in E. coli, Saccharomyces cerevisiae, and other unicellular organisms. In humans, defects in excision repair cause the inherited disease xeroderma pigmentosum (XP) (1). The mechanism of excision repair in S. cerevisiae, which is quite similar to human excision repair, has been reviewed elsewhere (2) and is not discussed here. Archaebacteria represent the third biological kingdom and DNA repair genes are also found in their genomes; surprisingly, excision repair proteins in Archaea may be either prokaryotic- or eukaryotic-like or a combination of the two (3, 4). E. coli-like excision of a 12–13 nucleotide-long damage-containing oligomer has been observed in Methanobacterium thermoautotrophicum (5); other species have not been tested for excision activity.

Ultraviolet (UV) radiation produces cyclobutane pyrimidine dimers and (6–4) photoproducts. Chemical agents that damage DNA are a structurally diverse group of compounds that bind DNA either directly or following metabolic activation to DNA reactive species. These agents include both known carcinogens [e.g., benzo(a)pyrene and acetylaminofluorene] and chemotherapeutic drugs (e.g., cisplatin, nitrogen mustard, and tamoxifen). Bifunctional chemical agents (e.g., psoralen, nitrogen mustard, and mitomycin C) can generate interstrand DNA cross-links that present a unique problem for the cell becuse both strands of the DNA molecule are damaged. Exogenous ionizing radiation and intracellular oxidative metabolism generate reactive oxygen species that damage bases (e.g., oxidized or reduced bases, ring-opening and fragmentation) and generate both single- and double-strand breaks in the DNA. DNA–protein cross-links may be generated by exogenous agents or during certain DNA transactions involving DNA-reactive intermediates. Mismatched bases occur at a low frequency during DNA replication and recombination or as a result of spontaneous or induced base deamination and are mutagenic if not corrected.

Under some circumstances cells do not (or cannot) repair DNA damage prior to the start of replication or cell division; in these cases the damage is tolerated. Tolerance mechanisms include the SOS response, translesion synthesis, and the damage checkpoint response. E. coli and other prokaryotes have a tightly regulated physiological response, called the SOS response, that permits cell survival in the presence of DNA damage (6). In the SOS response the expression of over 30 proteins, including a subset of repair proteins (UvrA, UvrB, UvrD), is induced following exposure to damaging agents, thus increasing the repair capacity. The upregulated proteins also include DNA polymerases responsible for lesion bypass (translesion synthesis), a mechanism that avoids replication arrest and cell death. However, translesion polymerases have reduced fidelity relative to replicative polymerases and cell survival is achieved at the cost of elevated mutagenesis; eukaryotes also have error-prone polymerases involved in translesion synthesis (7, 8).

There is no evidence for a true SOS response in eukaryotes, but there are suggestions that the DNA damage- and p53-dependent upregulation/stabilization of proteins may function as an SOS response (9). Eukaryotes do have a checkpoint response in which specific proteins sense the presence of DNA damage and signal the cell to stop the cell cycle, presumably allowing time for repair, before proceeding with replication or mitosis or, in the case of extensive damage or abnormally growing cells, the cells are sent to an apoptotic pathway. Mechanistic details of the checkpoint response are not well defined at present, but in humans the process requires damage sensors (e.g., ATM, ATR, Rad17-RFC, and the Rad9-Rad1-Hus1 complex), mediators (e.g., claspin, BRCA1, and MDC1), transducers (e.g., Chk1 and Chk2), and effectors (e.g., p53, Cdk2, and Cdc2) (10). It has been suggested that the bacterial SOS response constitutes a primitive DNA damage checkpoint (11).

Both prokaryotic and eukaryotic cells have multiple repair pathways to correct DNA damage or errors of misincorporation. These pathways may be elegantly simple, requiring a single protein, or exquisitely complex, involving multiple steps and the coordinated activity of many proteins. Repair is direct and error-free when the damage is corrected in situ, but repair involving incision of the DNA backbone to excise the damage results in a gapped DNA structure that is filled in by DNA polymerases; thus there is a potential for misincorporation and mutagenesis during excision repair processes. These repair pathways, including direct reversal of damage, base excision repair, and double-strand break repair, have been the subject of other reviews (10, 12, 13, 14, 15) and are not discussed here.

Nucleotide excision repair (excision repair) eliminates a broad spectrum of DNA damage by dual incisions bracketing the lesion. Damage is removed in the form of a 12–13 nucleotide-long oligomer in prokaryotes and in a 24–32 nucleotide-long oligomer in eukaryotes (16, 17) (Fig. 1). Excision repair is composed of three basic steps: (1) damage recognition, (2) dual incisions and release of the excised oligomer, and (3) resynthesis to fill in the gap and ligation (10, 18, 19). Nucleotide excision repair is the primary repair system for bulky DNA adducts such as the cyclobutane pyrimidine dimer (Pyr<>Pyr), (6–4) photoproduct, benzo(a)pyrene-guanine adduct, acetylaminofluorene-guanine (AAF-G), and cisplatin-d(GpG) diadduct. Because humans and other placental mammals lack photolyases, excision repair is the only known mechanism for the removal of sunlight-induced Pyr<>Pyr and (6-4) photoproducts, the most commonly encountered bulky DNA lesions. Additionally, evidence shows that nucleotide excision repair serves as a backup system for the repair of oxidatively damaged and alkylated bases as thymine glycol and 8-oxoguanine are also substrates for the E. coli and human excision nucleases (20, 21, 22).

In E. coli, dual incisions are accomplished by three proteins (UvrA, UvrB, and UvrC); in humans, 14–15 polypeptides in six repair factors carry out the same task. In contrast to all other repair systems, the prokaryotic and eukaryotic excision repair factors are not evolutionarily related and show no sequence homology to one another. However, the basic strategies for the prokaryotic and eukaryotic excision nucleases are similar and involve sequential steps coordinated via specific interactions between proteins and DNA. First, damage is recognized by an ATP-independent mechanism to form an unstable DNA–protein complex. Then this complex is converted to a stable preincision form by ATPase subunits that hydrolyze ATP and unwind the duplex to create an open structure and to promote formation of more intimate protein–DNA contacts. Finally, the dual incisions are made in a concerted, but asynchronous, manner such that the 3′ incision precedes the 5′ incision. Before presenting a detailed discussion of excision repair in E. coli and human cells, we address the major problem of damage recognition. This is the first and most crucial step in repair and yet, in excision repair, it is the least understood.

Section snippets

Damage Recognition

All repair mechanisms are initiated by the recognition of damaged or inappropriate bases in DNA or alterations in DNA structure. The biological dilemma in DNA repair is the virtually infinite number of DNA lesions that can and often do form within the life span of the cell and the necessity of repairing these lesions within a lifetime. In nature this problem has been solved by two main approaches. In one approach, exemplified by photolyase, a single enzyme with near-absolute specificity repairs

Excision Repair in Escherichia coli

Excision repair was discovered in E. coli as a system that removes UV-induced cyclobutane thymine dimers from DNA (53, 54), and it was determined that the excision of dimers required the products of three genes, UvrA, UvrB, and UvrC (54, 55). When E. coli excision repair was reconstituted in vitro with purified UvrA, UvrB, and UvrC, it was found that these three proteins are necessary and sufficient for making dual incisions at precise locations both 5′ and 3′ to the lesion, resulting in

Excision Repair in Humans

In humans, excision repair is the only known mechanism for the removal of UV-induced lesions. Therefore, we first present an overview of human diseases characterized by sensitivity to sunlight, the most common source of DNA damage for lesions introduced by exogenous agents. Then we summarize the structural features of excision repair proteins and known protein–protein interactions before discussing the mechanistic aspects of human excision repair. Finally, the section concludes with a brief

Modulation of DNA Repair by Transcription

Excision repair is affected by other DNA transactions, including binding of regulatory proteins, compaction into chromatin, replication, recombination, and transcription. It has been shown that transcription stimulates excision repair both in E. coli and in humans (235, 236, 237). In most cases, transcription stimulates the repair of only the transcribed strand (236) and it may actually inhibit repair of the transcribed strand in the absence of an active mechanism coupling the two processes (238

Repair of Chromatin

Eukaryotic chromosomes are packaged into chromatin, a compact structure comprised at the first level of repeating nucleosome cores tightly wrapped around a histone octamer and joined to linker DNA associated with a linker histone. This structural organization has a significant influence on the distribution of UV-induced damage within chromatin (nucleosome vs linker) and within the nucleosome core (258, 259). In addition to this effect on adduct distribution, packaging of DNA into nucleosomes

DNA Interstrand Cross-Links

Bifunctional chemical agents introduce interstrand cross-links (ICL), a unique problem to the cell because the DNA is damaged on both strands. ICL inducers include psoralen and chemotherapeutic agents such as mitomycin C and nitrogen mustards. ICLs are among the most toxic types of DNA damage because they inhibit strand separation and, therefore, interfere with replication, transcription, and chromosomal segregation. In E. coli, ICLs are repaired primarily through an error-free mechanism

Concluding Comments

The basic mechanism of excision repair in E. coli and humans is well understood. In general, prokaryotes and eukaryotes use similar strategies to recognize and remove DNA damage, but there are differences in mechanistic details and these differences are summarized here. (1) Damage recognition in E. coli is accomplished by the UvrA2B1 complex, with initial damage specificity provided by UvrA. In humans, three factors, XPA, RPA, and XPC (and, indirectly, TFIIH), are required for specific damage

References (295)

  • BranumM.E. et al.

    DNA repair excision nuclease attacks undamaged DNA: A potential source of spontaneous mutations

    J. Biol. Chem.

    (2001)
  • O'BrienP.J. et al.

    Dissecting the broad substrate specificity of human 3-methyladenine-DNA glycosylase

    J. Biol. Chem.

    (2004)
  • O'BrienP.J. et al.

    The Escherichia coli 3-methyladenine DNA glycosylase AlkA has a remarkably versatile active site

    J. Biol. Chem.

    (2004)
  • YeN. et al.

    Adaptive enhancement and kinetics of nucleotide excision repair in humans

    Mutat. Res.

    (1999)
  • HolmquistG.P.

    Endogenous lesions, S-phase-independent spontaneous mutations, and evolutionary strategies for base excision repair

    Mutat. Res.

    (1998)
  • HusainI. et al.

    Sequences of Escherichia coli uvrA gene and protein reveal two potential ATP binding sites

    J. Biol. Chem.

    (1986)
  • NavaratnamS. et al.

    Evidence from extended X-ray absorption fine structure and site-specific mutagenesis for zinc fingers in UvrA protein of Escherichia coli

    J. Biol. Chem.

    (1989)
  • ClaassenL.A. et al.

    Deletion mutagenesis of the Escherichia coli UvrA protein localizes domains for DNA binding, damage recognition, and protein-protein interactions

    J. Biol. Chem.

    (1991)
  • OhE.K. et al.

    Characterization of the helicase activity of the Escherichia coli UvrAB protein complex

    J. Biol. Chem.

    (1989)
  • HsuD.S. et al.

    Structure and function of the UvrB protein

    J. Biol. Chem.

    (1995)
  • SkorvagaM. et al.

    The β-hairpin motif of UvrB is essential for DNA binding, damage processing, and UvrC-mediated incisions

    J. Biol. Chem.

    (2002)
  • TangM.-S. et al.

    Two forms of UvrC protein with different double-stranded DNA binding affinities

    J. Biol. Chem.

    (2001)
  • Van HoutenB. et al.

    DNase I footprint of ABC excinuclease

    J. Biol. Chem.

    (1987)
  • Bertrand-BurggrafE. et al.

    Identification of the different intermediates in the interaction of (A)BC excinuclease with its substrates by Dnase I footprinting on two uniquely modified oligonucleotides

    J. Mol. Biol.

    (1991)
  • DelagoutteE. et al.

    Sequence-dependent modulation of nucleotide excision repair: The efficiency of the incision reaction is inversely correlated with the stability of the pre-incision UvrB-DNA complex

    J. Mol. Biol.

    (1997)
  • DelagoutteE. et al.

    The isomerization of the UvrB-DNA preincision complex couples the UvrB and UvrC activities

    J. Mol. Biol.

    (2002)
  • ShiQ. et al.

    Electron microscopic study of (A)BC excinuclease. DNA is sharply bent in the UvrB-DNA complex

    J. Mol. Biol.

    (1992)
  • LinJ.-J. et al.

    Active site of (A)BC excinuclease. II. Binding, bending, and catalysis mutants of UvrB reveal a direct role in 3′ and an indirect role in 5′ incision

    J. Biol. Chem.

    (1992)
  • GrossmanL. et al.

    Nucleotide excision repair, a tracking mechanism in search of damage

    J. Biol. Chem.

    (1993)
  • TheisK. et al.

    The nucleotide excision repair protein UvrB, a helicase-like enzyme with a catch

    Mutat. Res.

    (2000)
  • VerhoevenE.E.A. et al.

    Catalytic sites for 3′ and 5′ incision of Escherichia coli nucleotide excision repair are both located in UvrC

    J. Biol. Chem.

    (2000)
  • LinJ.-J. et al.

    Active site of (A)BC excinuclease. I. Evidence for 5′ incision by UvrC through a catalytic site involving Asp399, Asp438, Asp466, and His538 residues

    J. Biol. Chem.

    (1992)
  • PetitC. et al.

    Nucleotide excision repair: From E. coli to man

    Biochimie

    (1999)
  • OrrenD.K. et al.

    Formation and enzymatic properties of the UvrB•DNA complex

    J. Biol. Chem.

    (1990)
  • MoolenaarG.F. et al.

    The role of ATP binding and hydrolysis by UvrB during nucleotide excision repair

    J. Biol. Chem.

    (2000)
  • CleaverJ.E.

    Defective repair replication of DNA in xeroderma pigmentosum

    Nature

    (1968)
  • AravindL. et al.

    Conserved domains in DNA repair proteins and evolution of repair systems

    Nucleic Acids Res.

    (1999)
  • ÖgrünçM. et al.

    Nucleotide excision repair in the third kingdom

    J. Bacteriol.

    (1998)
  • WalkerG.C.

    Understanding the complexity of an organisms's responses to DNA damage

    Cold Spring Harbor Symp. Quant. Biol.

    (2000)
  • GoodmanM.F.

    Error-prone repair DNA polymerases in prokaryotes and eukaryotes

    Annu. Rev. Biochem.

    (2002)
  • PrakashS. et al.

    Translesion DNA synthesis in eukaryotes: A one- or two-polymerase affair

    Genes Dev.

    (2002)
  • HanawaltP.C.

    Subpathways of nucleotide excision repair and their regulation

    Oncogene

    (2002)
  • SancarA. et al.

    Molecular mechanisms of mammalian DNA repair and the DNA damage checkpoints

    Annu. Rev. Biochem.

    (2004)
  • SuttonM.D. et al.

    The SOS response: Recent insights into umuDC-dependent mutagenesis and DNA damage tolerance

    Annu. Rev. Gen.

    (2000)
  • SancarA.

    Structure and function of DNA photolyase and cryptochrome blue-light photoreceptors

    Chem. Rev.

    (2003)
  • McCulloughA.K. et al.

    Initiation of base excision repair: Glycosylase mechanisms and structures

    Annu. Rev. Biochem.

    (1999)
  • MolC.D. et al.

    DNA repair mechanisms for the recognition and removal of damaged DNA bases

    Annu. Rev. Biophys. Biomol. Struct.

    (1999)
  • WestS.C. et al.

    Double-strand break repair in human cells

    Cold Spring Harbor Symp. Quant. Biol.

    (2000)
  • HuangJ.-C. et al.

    Human nucleotide excision nuclease removes thymine dimers from DNA by incising the 22nd phosphodiester bond 5′ and the 6th phosphodiester bond 3′ to the photodimer

    Proc. Natl. Acad. Sci. USA

    (1992)
  • SancarA.

    DNA excision repair

    Annu. Rev. Biochem.

    (1996)
  • Cited by (257)

    View all citing articles on Scopus
    View full text