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Fidelity consequences of the impaired interaction between DNA polymerase epsilon and the GINS complex
•The dpb2-100 allele encodes the mutant form of Dpb2p, the noncatalytic subunit of Pol ɛ.•It does not change the biochemical properties of DNA Pol ɛ holoenzyme.•Dpb2-100p has impaired interaction with DNA Pol ɛ catalytic subunit and GINS complex.•We report an increased participation of error-prone D...
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Published in: | DNA repair 2015-05, Vol.29, p.23-35 |
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Main Authors: | , , , , , , , |
Format: | Article |
Language: | English |
Subjects: | |
Citations: | Items that this one cites Items that cite this one |
Online Access: | Get full text |
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Summary: | •The dpb2-100 allele encodes the mutant form of Dpb2p, the noncatalytic subunit of Pol ɛ.•It does not change the biochemical properties of DNA Pol ɛ holoenzyme.•Dpb2-100p has impaired interaction with DNA Pol ɛ catalytic subunit and GINS complex.•We report an increased participation of error-prone DNA Pol ζ in this mutator strain.•We suggest involvement of Pol δ in the leading strand replication in dpb2-100 mutant.
DNA polymerase epsilon interacts with the CMG (Cdc45-MCM-GINS) complex by Dpb2p, the non-catalytic subunit of DNA polymerase epsilon. It is postulated that CMG is responsible for targeting of Pol ɛ to the leading strand. We isolated a mutator dpb2-100 allele which encodes the mutant form of Dpb2p. We showed previously that Dpb2-100p has impaired interactions with Pol2p, the catalytic subunit of Pol ɛ. Here, we present that Dpb2-100p has strongly impaired interaction with the Psf1 and Psf3 subunits of the GINS complex. Our in vitro results suggest that while dpb2-100 does not alter Pol ɛ’s biochemical properties including catalytic efficiency, processivity or proofreading activity – it moderately decreases the fidelity of DNA synthesis. As the in vitro results did not explain the strong in vivo mutator effect of the dpb2-100 allele we analyzed the mutation spectrum in vivo. The analysis of the mutation rates in the dpb2-100 mutant indicated an increased participation of the error-prone DNA polymerase zeta in replication. However, even in the absence of Pol ζ activity the presence of the dpb2-100 allele was mutagenic, indicating that a significant part of mutagenesis is Pol ζ-independent. A strong synergistic mutator effect observed for transversions in the triple mutant dpb2-100 pol2-4 rev3Δ as compared to pol2-4 rev3Δ and dpb2-100 rev3Δ suggests that in the presence of the dpb2-100 allele the number of replication errors is enhanced. We hypothesize that in the dpb2-100 strain, where the interaction between Pol ɛ and GINS is weakened, the access of Pol δ to the leading strand may be increased. The increased participation of Pol δ on the leading strand in the dpb2-100 mutant may explain the synergistic mutator effect observed in the dpb2-100 pol3-5DV double mutant. |
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ISSN: | 1568-7864 1568-7856 |
DOI: | 10.1016/j.dnarep.2015.02.007 |