Eukaryotic polymerases ι and ζ act sequentially to bypass DNA lesions

Abstract

DNA lesions can often block DNA replication, so cells possess specialized low-fidelity, and often error-prone, DNA polymerases that can bypass such lesions and promote replication of damaged DNA1. The Saccharomyces cerevisiae RAD30 and human hRAD30A encode Polη, which bypasses a cis–syn thymine–thymine dimer efficiently and accurately2,3,4,5,6,7. Here we show that a related human gene, hRAD30B8, encodes the DNA polymerase Polι, which misincorporates deoxynucleotides at a high rate. To bypass damage, Polι specifically incorporates deoxynucleotides opposite highly distorting or non-instructional DNA lesions. This action is combined with that of DNA polymerase Polζ, which is essential for damage-induced mutagenesis, to complete the lesion bypass. Polζ is very inefficient in inserting deoxynucleotides opposite DNA lesions, but readily extends from such deoxynucleotides once they have been inserted. Thus, in a new model for mutagenic bypass of DNA lesions in eukaryotes, the two DNA polymerases act sequentially: Polι incorporates deoxynucleotides opposite DNA lesions, and Polζ functions as a mispair extender.

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Figure 1: Purification and activity of human Rad30B protein.
Figure 2: Deoxynucleotide incorporation by Polι on undamaged and damaged DNA and its role in damage bypass.
Figure 3: Comparison of Polζ mispair extension and mispair insertion efficiencies.

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Acknowledgements

We thank D. Hinkle for plasmids pGST-Rev3 and pRev7 and R. Hodge for providing the T–T dimer- and (6-4) photoproduct-containing DNAs, the construction of which was supported by a National Institute of Environmental Health Science (NIEHS) Centre Grant. This work was supported by a grant from the NIH.

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Sealy Centre for Molecular Science, University of Texas Medical Branch at Galveston, 6.104 Medical Research Building, 11th and Mechanic Streets, Galveston, Texas 77555-1061, USA

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Johnson, R., Washington, M., Haracska, L. et al. Eukaryotic polymerases ι and ζ act sequentially to bypass DNA lesions. Nature 406, 1015–1019 (2000). https://doi.org/10.1038/35023030

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