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Published on: February 25, 2017
Low fidelity DNA synthesis by human DNA polymerase-eta
T Matsuda1, K Bebenek, C Masutani
1Laboratory of Molecular Genetics, National Institute of Environmental Health Sciences, Research Triangle Park, North Carolina 27709, USA.
Abstract:
A superfamily of DNA polymerases that bypass lesions in DNA has been described. Some family members are described as error-prone because mutations that inactivate the polymerase reduce damage-induced mutagenesis. In contrast, mutations in the skin cancer susceptibility gene XPV, which encodes DNA polymerase (pol)-eta, lead to increased ultraviolet-induced mutagenesis. This, and the fact that pol-eta primarily inserts adenines during efficient bypass of thymine-thymine dimers in vitro, has led to the description of pol-eta as error-free. However, here we show that human pol-eta copies undamaged DNA with much lower fidelity than any other template-dependent DNA polymerase studied. Pol-eta lacks an intrinsic proofreading exonuclease activity and, depending on the mismatch, makes one base substitution error for every 18 to 380 nucleotides synthesized. This very low fidelity indicates a relaxed requirement for correct base pairing geometry and indicates that the function of pol-eta may be tightly controlled to prevent potentially mutagenic DNA synthesis.
Insights
Human DNA polymerase eta (pol-eta), previously thought error-free, exhibits low fidelity when copying undamaged DNA. This DNA polymerase lacks proofreading, leading to significant base substitution errors and suggesting its function requires tight control.
Area of Science:
- Molecular Biology
- Genetics
- Biochemistry
Background:
- DNA polymerases are crucial for DNA replication and repair.
- Lesion-bypass DNA polymerases are a specialized family involved in DNA damage tolerance.
- Human DNA polymerase eta (pol-eta) is encoded by the XPV gene, linked to skin cancer susceptibility.
Purpose of the Study:
- To investigate the fidelity of human DNA polymerase eta (pol-eta) during replication of undamaged DNA.
- To determine if pol-eta's previously described 'error-free' bypass of thymine-thymine dimers accurately reflects its overall fidelity.
Main Methods:
- In vitro synthesis assays using purified human pol-eta.
- Analysis of base substitution error rates on undamaged DNA templates.
- Assessment of intrinsic proofreading exonuclease activity in pol-eta.
Main Results:
- Human pol-eta exhibits significantly lower fidelity on undamaged DNA compared to other template-dependent DNA polymerases.
- Pol-eta lacks intrinsic proofreading exonuclease activity.
- Error rates range from one base substitution per 18 to 380 nucleotides synthesized, depending on the mismatch.
Conclusions:
- Human pol-eta is not error-free and synthesizes undamaged DNA with very low fidelity.
- The relaxed base-pairing requirements suggest pol-eta's function may be tightly regulated to prevent mutagenic synthesis.
- Findings challenge the 'error-free' designation and highlight potential risks of uncontrolled pol-eta activity.
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