Abstract
Sulfolobus islandicus codes for four DNA polymerases: three are of the B-family (Dpo1, Dpo2, and Dpo3), and one is of the Y-family (Dpo4). Western analysis revealed that among the four polymerases, only Dpo2 exhibited DNA damage-inducible expression. To investigate how these DNA polymerases could contribute to DNA damage tolerance in S. islandicus, we conducted genetic analysis of their encoding genes in this archaeon. Plasmid-borne gene expression revealed that Dpo2 increases cell survival upon DNA damage at the expense of mutagenesis. Gene deletion studies showed although dpo1 is essential, the remaining three genes are dispensable. Furthermore, although Dpo4 functions in housekeeping translesion DNA synthesis (TLS), Dpo2, a B-family DNA polymerase once predicted to be inactive, functions as a damage-inducible TLS enzyme solely responsible for targeted mutagenesis, facilitating GC to AT/TA conversions in the process. Together, our data indicate that Dpo2 is the main DNA polymerase responsible for DNA damage tolerance and is the primary source of targeted mutagenesis. Given that crenarchaea encoding a Dpo2 also have a low-GC composition genome, the Dpo2-dependent DNA repair pathway may be conserved in this archaeal lineage.
Highlights
Multiple DNA polymerases are required in all organisms to duplicate their genomes and repair DNA lesions generated from endogenous compounds and environmental factors
We found that (a) no apparent fluctuation of protein content was observed for Dpo1, Dpo3, and Dpo4 during the drug treatment and that (b) the Dpo2 protein, present at a level below the detection limit before the drug addition, attained the highest level at 6 h post NQO addition (p.n.a.), with the plateau level maintained for 9 h in the cell before dropping down, and became barely detectable at 24 h p.n.a. (Figure 1)
These results suggested that the content of Dpo2 could be correlated to the level of DNA damage in S. islandicus
Summary
Multiple DNA polymerases are required in all organisms to duplicate their genomes and repair DNA lesions generated from endogenous compounds and environmental factors. Whereas Escherichia coli, the best-studied bacterial model, contains five DNA polymerases, the human genome codes for >15 such enzymes (Vaisman and Woodgate, 2017). These DNA polymerases are of two main types based on their cellular functions: those that are devoted to copy genomes with high fidelity and processivity (replicative polymerases) and those that mainly function in DNA repair (specialized polymerases) (McCulloch and Kunkel, 2008). Four families of replicative enzymes are known, that is, Families A, B, C, and D These replicative polymerases are endowed with the 3 –5 exonuclease activity that proofreads the synthesized product, and their active centers are only compatible with the Watson–Crick base paring, a characteristic that is essential for copying DNA with high accuracy. Most specialized polymerases capable of translesion DNA synthesis (TLS) belong to the Y-family, and they bypass DNA lesions by accommodating bulky or distorted template
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