Annu Rev Biochem 2002 71 133 63 DOI 10 1146 annurev biochem 71 090501 150041 Copyright 2002 by Annual Reviews All rights reserved First published as a Review in Advance on March 15 2002 EUKARYOTIC DNA POLYMERASES Ulrich Hu bscher 1 Giovanni Maga 1 2 and Silvio Spadari2 1 Institute of Veterinary Biochemistry and Molecular Biology University of Zu rich Winterthurerstrasse 190 CH 8057 Zu rich Switzerland e mail hubscher vetbio unizh ch 2 Istituto di Genetica Biochimica ed Evoluzionistica IGBE CNR Via Abbiategrasso 207 I 27100 Pavia Italy e mail maga igbe pv cnr it spadari igbe pv cnr it Key Words replication repair recombination translesion synthesis holoenzyme clamp clamp loaders f Abstract Any living cell is faced with the fundamental task of keeping the genome intact in order to develop in an organized manner to function in a complex environment to divide at the right time and to die when it is appropriate To achieve this goal an efficient machinery is required to maintain the genetic information encoded in DNA during cell division DNA repair DNA recombination and the bypassing of damage in DNA DNA polymerases pols and are the key enzymes required to maintain the integrity of the genome under all these circumstances In the last few years the number of known pols including terminal transferase and telomerase has increased to at least 19 A particular pol might have more than one functional task in a cell and a particular DNA synthetic event may require more than one pol which suggests that nature has provided various safety mechanisms This multi functional feature is especially valid for the variety of novel pols identified in the last three years These are the lesion replicating enzymes pol pol pol pol and Rev1 and a group of pols called pol pol pol pol and pol that fulfill a variety of other tasks CONTENTS INTRODUCTION THE CLASSICAL DNA POLYMERASES AND An Evolutionary Perspective DNA Polymerases Have a Very Conserved Active Site A Mechanistic Perspective DNA Polymerases Are Built by Addition of Specific Domains to a Conserved Core with Essential Catalytic Activity Coordinated Leading and Lagging Strand Synthesis and the DNA Polymerase Switch Mechanism Distinct Roles for DNA Polymerases and Fidelity of DNA Synthesis Novel Roles for Accessory Proteins The Matchmaker Concept for Establishing a Moving Platform Replication Factor C and Proliferating Cell Nuclear Antigen DNA Polymerase The Mitochondrial Replicase 0066 4154 02 0707 0133 14 00 134 136 136 138 141 144 144 146 133 134 HU BSCHER y MAGA y SPADARI Further Functions of DNA Polymerases Primase and A Coordinated Interplay DNA Polymerase The Prototype of a Repair Enzyme THE NOVEL DNA POLYMERASES Discovery Functions of DNA Polymerases and Rev1 The Lesion Replicating Enzymes How Are the Functions of Lesion Bypassing DNA Polymerases Coordinated DNA Polymerases and Terminal Deoxynucleotidyl Transferase Enzymes with Further Distinct Functions How Many DNA Polymerases Are Involved in the Immune System FUTURE DIRECTIONS 147 150 151 151 151 156 156 157 158 INTRODUCTION Complex cellular functions are performed by networks of protein machines These protein assemblies contain highly coordinated moving parts whose functions are in general temporally and spatially regulated by a series of ordered conformational changes that and are powered by chemical energy derived from hydrolysis of nucleoside triphosphates 1 The eukaryotic replisome is an example of how protein components interact and communicate with one another acting in a coordinated fashion in order to duplicate the genetic information of the cell 2 At the heart of the replisome are template directed machines for phosphoryl transfer 3 the DNA polymerases pols Since the discovery of pol in 1957 the number of eukaryotic pols identified has grown reviewed in 4 In the early 1970s pols and were discovered leading to the simple concept that pol was the enzyme responsible for nuclear DNA replication pol for DNA repair and pol for mitochondrial DNA replication Table 1 The discovery of pol and pol and the intensive work done on them during the 1980s suggested that a particular pol might have more than one functional task and that a particular DNA synthetic event may require more than one pol reviewed in 5 Moreover since 1999 at least 10 novel pols have been discovered for details see below Since both DNA replication and repair are of primary importance for cells it appears that nature created safety mechanisms by employing different pols for similar functional tasks For example DNA replication requires pol pol and pol while translesion DNA synthesis depends at least on pol pol pol pol and Rev1 6 In many cases pols are multipolypeptide complexes that contain other functional subunits in addition to the polymerizing subunit which often displays a proofreading 3 3 5 exonuclease The other functional subunits are responsible for other enzymatic activities e g DNA primase to synthesize RNA primers or allow the pol to interact with other proteins An impressive example of these multiple functions is the finding that the replicative pols and are chaperoned by two accessory proteins replication factor C and proliferating cell nuclear antigen reviewed in 7 and 8 respectively which allow accurate and fast DNA synthesis At EUKARYOTIC DNA POLYMERASES 135 TABLE 1 The classical DNA polymerases and telomerasea Pol Functional tasks Initiator pol b POLA B Lagging strand pol c Base excision repair pol POLB Recombination pol X Meiosis pol Translesion pol Role in neurogenesis Mitochondrial replication pol POLG A Main pol at the leading and lagging strand POLD Base excision repair pol B Nucleotide excision repair pol Mismatch repair pol Double strand break repair pol Recombination pol Leading and lagging strand pol POLE Base excision repair pol B Recombination pol Checkpoint control pol Telomerase Telomere maintenance pol Homologous to reverse transcriptase a For details see text and references therein b c Human Genome Organization HUGO nomenclature DNA polymerase family A B X or Y see References 10 and 11 the structural level all classical pols share a similar active site 9 Polymerization occurs through a mechanism catalyzed by two metal ions this mechanism guarantees the incorporation of the correctly base paired deoxyribonucleoside monophosphate onto a growing primer template duplex with the exception of some of the translesion pols e g pol 6 This review first summarizes the assembled knowledge of the five