Translesion Synthesis of 2′-Deoxyguanosine Lesions by Eukaryotic DNA Polymerases

Abstract

With the discovery of translesion synthesis DNA polymerases, great strides have been made in the last two decades in understanding the mode of replication of various DNA lesions in prokaryotes and eukaryotes. A database search indicated that approximately 2000 articles on this topic have been published in this period. This includes research involving genetic and structural studies as well as in vitro experiments using purified DNA polymerases and accessory proteins. It is a daunting task to comprehend this exciting and rapidly emerging area of research. Even so, as the majority of DNA damage occurs at 2′-deoxyguanosine residues, this perspective attempts to summarize a subset of this field, focusing on the most relevant eukaryotic DNA polymerases responsible for their bypass.

Mammalian DNA Polymerase Kappa Activity and Specificity

“…Pol κ is inhibited by the major groove adducts BPDE- N 6 -dA [14], N 6 -furfuryl-dA [48], and bulky O 6 -alkyl-dG modifications [49,50]. Pol κ can replicate DNA templates containing various bulky and non-bulky lesions, such as BPDE- N 2 -dG [10,14,16,17,51,52,53,54,55], N 2 -(1-carboxyethyl)-dG [56,57], 8-oxo-dG [17], N 2 -alkyl-dG [10,58], O 2 - and some O 4 -alkyl-dT adducts [59,60,61,62], thymine glycol [45], DNA-peptide crosslinks, and intrastrand adducts and interstrand crosslinks (ICL) formed between purine bases and cisplatin [6,10,17,63,64,65,66,67,68,69,70].…”

“…Benzo[a]pyrene. One of the most well-characterized bulky adducts, 10S- trans - anti -benzo[a]pyrene-7,8-dihydrodiol-9,10-epoxide (BPDE- N 2 -dG) (Figure 3), is derived from an environmental mutagen, B[a]P, present in tobacco smoke and combustion products of fossil fuel [65]. Replicative polymerases stall at BPDE- N 2 -dG, which is predominantly repaired through nucleotide excision repair (NER).…”

Error-Prone and Error-Free Translesion DNA Synthesis over Site-Specifically Created DNA Adducts of Aryl Hydrocarbons (3-Nitrobenzanthrone and 4-Aminobiphenyl)

“…To avoid cell death, cells have mechanisms to complete DNA replication by bypassing these adducts. There are two mechanisms for DNA replication bypass: one is damage avoidance including replication fork regression and recombination repair, which are error-free mechanisms; the other is TLS, which is either error-free or error-prone depending on the adduct structure and the DNA polymerases involved ( 19 , 20 ). The DNA polymerases participating in TLS are translesion DNA polymerases, which are different from the replicative DNA polymerases δ and ɛ.…”

“…The major translesion DNA polymerases are Pol η, κ, ι, and ζ and Rev1 ( 21 , 22 ). When a DNA replication fork is stalled at a DNA adduct, mono-ubiquitination of proliferative cell nuclear antigen (PCNA) by Rad6/Rad18 proteins triggers the replacement of Pol δ or ɛ with Pol η, κ, ι, or ζ or Rev1, which continues DNA synthesis over the adduct ( 19 , 20 ). Given that translesion DNA polymerases are deficient in 3 ′ to 5 ′ exonuclease activity (proofreading activity), these polymerases readily insert an incorrect nucleotide opposite the adducted nucleotide and cause a base-change mutation.…”

EGFP Reporters for Direct and Sensitive Detection of Mutagenic Bypass of DNA Lesions

“…Most of the relevant knowledge has come from artificially constructed TLS templates, which were delivered to suitable host cells for accomplishment of DNA synthesis over the damaged nucleotide. Thus, vectors carrying defined synthetic DNA modifications were designed either for integration into chromosomal DNA [3] or, more commonly, for allowing extrachromosomal TLS and repair followed by shuttling to bacteria and sequencing of mutations in the survived fraction of transformation-competent vector DNA [13][14][15][16]. The efficiency of detection of both faithful and mutagenic TLS can be immensely enhanced by accommodating the DNA lesion within a reporter gene [17,18]; however, for the fidelity assessment, TLS reporters described in the literature also require shuttling into bacteria [19].…”