1. Biochemistry and Chemical Biology
  2. Chromosomes and Gene Expression
Download icon

Mutations: The dark side of DNA repair

  1. Samuel H Wilson  Is a corresponding author
  1. National Institutes of Health, United States
  • Cited 5
  • Views 2,340
  • Annotations
Cite this article as: eLife 2014;3:e03068 doi: 10.7554/eLife.03068


The BER pathway is widely used to repair DNA damage in cells, but it can also introduce unwanted mutations and is sometimes hijacked by other pathways.

Main text

DNA can be damaged in many ways—bases can, for example, be oxidized, alkylated or deaminated—so cells need a way to repair DNA damage. Sometimes it is possible to reverse the process that caused these ‘base lesions’, but cells tend to employ a more complex approach that is called the base excision repair (BER) pathway. This process involves three basic steps: first, enzymes called nucleases break the DNA strand that is damaged on either side of the lesion so that the section containing the lesion can be removed; second, DNA polymerases make a new stretch of DNA to replace the section that has been removed, using the undamaged strand as a template; third, DNA ligases join this new stretch to the existing DNA.

However, there is a ‘dark side’ to DNA repair because the process by which the replacement DNA is synthesized is inherently error-prone. Moreover, DNA is normally tightly packaged inside the nucleus to protect it from damage: however, the BER pathway involves regions of DNA being unpackaged, and this provides other enzymes and repair pathways with access to the DNA, which can lead to lesions and mutations. The overall result is that DNA repair can sometimes lead to more rather than less damage and, in some cases, this damage can ultimately kill the cell (Fu et al., 2012).

Large-scale DNA sequencing has revealed a complex pattern of lesions and mutations that includes mutational hotspots and mutations that are linked to specific sequences of bases. In some cases the mutations appear to be linked to enzymes belonging to the APOBEC family—which change the cytosine bases in DNA into uracil bases—being able to access the DNA (Roberts et al., 2013; Nik-Zainal et al., 2014). Finally, some seemingly random mutations occur with high frequency.

Now, in eLife, Jia Chen, Brendan Miller and Anthony Furano—who all are based at the National Institute of Diabetes and Digestive and Kidney Diseases—have used DNA sequencing to explore the accuracy of the BER pathway in mammalian cells (Chen et al., 2014). They used genetic techniques to deliberately introduce lesions associated with different sub-pathways of the BER pathway. These lesions included: a simple strand break between two neighbouring bases (which is known as a nick); replacing the cytosine base opposite a guanine base with an abasic site; and various types of base mismatches (such as thymine/guanine, uracil/guanine or hydroxymethyl-uracil/guanine instead of cytosine/guanine). The lesions were individually engineered into episomal DNA, and the episomes were allowed to undergo repair in mammalian cells and then isolated. After passage through E. coli, mutants were selected, and the DNA was subjected to sequence analysis.

As expected, most of the lesions were repaired by the BER pathway (Figure 1), but there were interesting differences in the repair of the different types of lesions. In some cases, for example, there were mutations in the template strand, rather than the strand containing the lesion, which suggests that various enzymes had been able to access the template strand during the repair process.

The BER pathway can repair DNA damage, but it can also induce mutations.

Cells rely on the base excision repair (BER) pathway to repair base lesions in DNA. A single lesion (X) in one of the strands in double-stranded DNA can be repaired by removing the damaged base (and some neighbouring bases) and replacing it with a new stretch of DNA (shown by a rectangle): this process is called long-patch BER (left). However, the process used to synthesize the new DNA is prone to error, which can lead to the introduction of mutations (symbol). FEN1 is the nuclease that removes the flap of single-stranded DNA (ssDNA); Pol ß is the polymerase that synthesises the new DNA. It is also possible to replace just the damaged base (middle), but this single-nucleotide BER pathway can also introduce mutations. Sometimes, however, the BER pathway is hijacked by a DNA repair pathway called the mismatch repair (MMR) system that exposes a stretch of ssDNA on the 5′side of the lesion: this ssDNA then becomes a substrate for APOBEC enzymes that remove amine groups from cytosine bases, which introduces even more mutations (right). Note: figure not drawn to scale; the orientation of strands is shown.

Moreover, mutations were observed in the bases on one side of the base lesion (distal to the lesion). These ‘flanking mutations’ were shown to depend on the initial steps of the BER pathway in most cases. A number of enzymes are known to be capable of exposing single-stranded DNA on either side of a nick, but it seems surprising that the molecular machinery responsible for the BER pathway allowed these enzymes to have access to the lesions. Previously it was thought that each enzyme in the BER pathway ‘handed over’ to the next enzyme in a highly coordinated process that left little scope for third parties to have access to the DNA (Prasad et al., 2010), but the presence of the flanking mutations suggests that the lesion recognition and hand-over process is not always able to keep third parties away from the lesion.

So what actually causes the flanking mutations? Chen, Miller and Furano suggest that the BER pathway is ‘hijacked’ by another DNA repair pathway called the mismatch repair system (which, as its name suggests, repairs base mismatches). They provided evidence for this by showing that depleting factors involved in the mismatch repair system results in a reduction in flanking mutations.

Chen et al. also noticed that many of the flanking mutations involved cytosine bases that were preceded by thymine bases, with the cytosine bases changing to uracil bases. This pattern suggested that certain APOBEC enzymes—those that have a preference for the TpC sequence in single-stranded DNA—were involved. To test this idea Chen et al. depleted the level of APOBEC enzymes and, as they predicted, there was a drop in the number of flanking mutations. Taken together, the results point to the idea that the mismatch repair machinery exposes single-stranded DNA that serves as a substrate for the APOBEC enzymes, which remove amine groups from the cytosine bases to leave uracil bases (Roberts et al., 2012, 2013). DNA synthesis to copy the single-stranded DNA then leads to mutations.

Overall, the results reported by Chen, Miller and Furano open up a whole new range of questions about the introduction of mutations during DNA repair.


Article and author information

Author details

  1. Samuel H Wilson

    Laboratory of Structural Biology, National Institute of Environmental Health Sciences, National Institutes of Health, Research Triangle Park, United States
    For correspondence
    Competing interests
    The author declares that no competing interests exist.

Publication history

  1. Version of Record published: May 20, 2014 (version 1)


© 2014, Wilson

This article is distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use and redistribution provided that the original author and source are credited.


  • 2,340
    Page views
  • 81
  • 5

Article citation count generated by polling the highest count across the following sources: Crossref, Scopus, PubMed Central.

Download links

A two-part list of links to download the article, or parts of the article, in various formats.

Downloads (link to download the article as PDF)

Download citations (links to download the citations from this article in formats compatible with various reference manager tools)

Open citations (links to open the citations from this article in various online reference manager services)

Further reading

    1. Biochemistry and Chemical Biology
    2. Cancer Biology
    Regina L Agulto et al.
    Research Article

    The microtubule-associated protein, doublecortin-like kinase 1 (DCLK1), is highly expressed in a range of cancers and is a prominent therapeutic target for kinase inhibitors. The physiological roles of DCLK1 kinase activity and how it is regulated remain elusive. Here, we analyze the role of mammalian DCLK1 kinase activity in regulating microtubule binding. We find that DCLK1 autophosphorylates a residue within its C-terminal tail to restrict its kinase activity and prevent aberrant hyperphosphorylation within its microtubule-binding domain. Removal of the C-terminal tail or mutation of this residue causes an increase in phosphorylation within the doublecortin domains, which abolishes microtubule binding. Therefore, autophosphorylation at specific sites within DCLK1 have diametric effects on the molecule's association with microtubules. Our results suggest a mechanism by which DCLK1 modulates its kinase activity to tune its microtubule-binding affinity. These results provide molecular insights for future therapeutic efforts related to DCLK1's role in cancer development and progression.

    1. Biochemistry and Chemical Biology
    2. Cell Biology
    Janani Gopalan et al.
    Research Article Updated

    Pathophysiological defects in water homeostasis can lead to renal failure. Likewise, common genetic disorders associated with abnormal cytoskeletal dynamics in the kidney collecting ducts and perturbed calcium and cAMP signaling in the ciliary compartment contribute to chronic kidney failure. We show that collecting ducts in mice lacking the A-Kinase anchoring protein AKAP220 exhibit enhanced development of primary cilia. Mechanistic studies reveal that AKAP220-associated protein phosphatase 1 (PP1) mediates this phenotype by promoting changes in the stability of histone deacetylase 6 (HDAC6) with concomitant defects in actin dynamics. This proceeds through a previously unrecognized adaptor function for PP1 as all ciliogenesis and cytoskeletal phenotypes are recapitulated in mIMCD3 knock-in cells expressing a phosphatase-targeting defective AKAP220-ΔPP1 mutant. Pharmacological blocking of local HDAC6 activity alters cilia development and reduces cystogenesis in kidney-on-chip and organoid models. These findings identify the AKAP220-PPI-HDAC6 pathway as a key effector in primary cilia development.