Roles of DNA Polymerases in Replication, Repair, and Recombination in Eukaryotes

Youri I. Pavlov*,†; Polina V. Shcherbakova*,‡; Igor B. Rogozin§,¶    * Eppley Institute for Research in Cancer and Allied Diseases
† Departments of Biochemistry and Molecular Biology, and Pathology and Microbiology, University of Nebraska Medical Center, Omaha, Nebraska 68198-6805
‡ Department of Genetics, Cell Biology and Anatomy, University of Nebraska Medical Center, Omaha, Nebraska 68198-6805
§ National Center for Biotechnology Information, National Library of Medicine, National Institutes of Health, Bethesda, Maryland 20894
¶ Institute of Cytology and Genetics, Novosibirsk 630090, Russia

I Introduction

DNA replication is one of the most fundamental processes in biology. It is required for the proper transmission of genetic information. Accurate and efficient replication and repair of genomic DNA are the bases for the evolutionary determined level of conservation of the genetic information and the prevention of genetic diseases (Bielas and Loeb, 2005; Friedberg et al., 2002; Kondo, 1973; Loeb et al., 1974; Radman, 1999).

The mechanism of replication is a polynucleotide template-directed polymerization of deoxynucleoside triphosphates by a DNA polymerase using a “two-metal-ion” mechanism (Kornberg and Baker, 1991; Steitz, 1998). The synthesis occurs exclusively in a 5′ to 3′ direction. Therefore, the two antiparallel strands of DNA duplexes should be replicated by somewhat different machineries (Garg and Burgers, 2005b; McHenry, 2003). The current model postulates that the leading strand is synthesized continuously, whereas the lagging strand is synthesized in short patches that are sealed together by DNA ligase (Garg and Burgers, 2005b; Johnson and O'Donnell, 2005).

In addition to the replication of undamaged DNA, DNA polymerases are also involved in the replication and repair of damaged DNA. They participate in various excision repair pathways, in recombination repair, or in bypassing the blocking adducts, thus forming a network of proteins that acts sequentially in the maintenance of genome integrity (Budd et al., 2005; Stauffer and Chazin, 2004). A wide diversity of DNA substrates in various DNA transactions are used by DNA polymerases belonging to several structural families. This review summarizes the current knowledge about the roles of different DNA polymerases in DNA replication, repair, and recombination. We focus on DNA template-dependent DNA polymerases, omitting terminal transferases, reverse transcriptases, telomerases, and RNA polymerases, which are reviewed extensively elsewhere (Benedict et al., 2000; Boeger et al., 2005; Collins, 1996; Cramer, 2004; Kelleher et al., 2002; Lingner and Cech, 1998; Ren and Stammers, 2005).

Here we are unable to cover the immerse literature in the field and will focus on the main functions of DNA polymerases. During the past 3 years, excellent reviews on various aspects of DNA polymerases appeared: on replisome assembly (Johnson and O'Donnell, 2005), on polymerases at the fork (Garg and Burgers, 2005b), on the general functions of DNA polymerases (Bebenek and Kunkel, 2004) and their mechanisms related to structure (Rothwell and Waksman, 2005), on the mechanisms of translesion DNA synthesis (Prakash et al., 2005), on the structure of translesion DNA polymerases (Yang, 2005), on the mechanisms of polymerase switch (Friedberg, 2005; Friedberg et al., 2005; Ulrich, 2005b), on DNA polymerase fidelity (Kunkel, 2004), on the regulation of DNA replication through the S phase (Takeda and Dutta, 2005), on replication complexes in genome stability (Toueille and Hubscher, 2004), and many others. We cite only cornerstone experimental papers and most frequently refer the reader to recent reviews.

II Classification and the Main Properties of Eukaryotic DNA Polymerases

A Overview of the Maintenance of Genome Stability

The bases in cellular DNA are continuously damaged by spontaneous hydrolysis and oxidation and other endogenous and environmental mutagens (Table I). The mutation rate in mammals, however, is kept low with one mutation or less per effective genome per sexual generation (Drake, 1999). Accurate and efficient replication and repair processes have evolved to achieve this goal. Most of these processes include DNA synthesis by DNA polymerases (Fig. 1). Intact DNA molecules are replicated with high fidelity (up to 10−11 per base replicated) due to three sequential fidelity control steps: base selection by DNA polymerases, exonucleolytic proofreading, and DNA mismatch repair (MMR) (Fig. 1, pathway A) (Kunkel and Bebenek, 2000; Morrison et al.,1993; Schaaper, 1993). It is important for accurate replication that the deoxyribonucleoside triphosphate (dNTP) pools are devoid of mutagenic contaminants (Maki and Sekiguchi, 1992) and balanced in a way to ensure the best performance of fidelity mechanisms (Chabes and Thelander, 2003; Kunz, 1988; Mathews and Ji, 1992; Meuth, 1989). Damaged nucleotides in DNA can be repaired by nucleotide excision repair (NER) or base excision repair (BER) reactions (left part of Fig. 1). In this case, a polymerase uses the intact information of the undamaged DNA strand to restore the correct nucleotide sequence. Intermediates of NER and BER are single-strand breaks that may be converted to double-strand breaks (DSB) during replication (Courcelle and Hanawalt, 2003; Hanawalt, 1966; Kouzminova and Kuzminov, 2006). DSB can also be induced by a variety of genotoxicants (Friedberg et al., 2006). The breaks are extremely dangerous unless repaired by either homologous recombination using an intact homologous duplex or by nonhomologous end-joining (NHEJ) that might require specialized DNA polymerases for processing DNA ends. The damaged templates could also be replicated by specialized translesion synthesis (TLS) DNA polymerases whose active sites can accommodate unusual base pairs (DNA damage bypass, pathway B in Fig. 1). Some agents cause cross-links between antiparallel DNA strands. In this case, the genetic information at the damaged site is lost and can be restored only by recombination with an intact homologous DNA molecule. The pathway of cross-link repair involves a combination of NER, TLS, and recombination. In addition to the replication, repair, and recombination pathways that prevent genome instability, DNA polymerases can also participate in the developmental processes that require elevated mutagenesis in a small part of the genome. For example, to generate immunoglobulin variability, DNA damage is introduced by specific enzymes into certain genomic sites and is repaired by error-prone processes (Section IV.E).