Fidelity consequences of the impaired interaction between DNA polymerase epsilon and the GINS complex

DNA polymerase epsilon interacts with the CMG (Cdc45-MCM-GINS) complex by Dpb2p, the noncatalytic subunit of DNA polymerase epsilon. It is postulated that CMG is responsible for targeting of Pot epsilon 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 epsilon. Here, we present that Dpb2-100p has strongly impaired interaction with the Psfl and Psf3 subunits of the GINS complex. Our in vitro results suggest that while dpb2-100 does not alter Pol epsilon'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 zeta activity the presence of the dpb2-100 allele was mutagenic, indicating that a significant part of mutagenesis is Pot zeta-independent. A strong synergistic mutator effect observed for transversions in the triple mutant dpb2-100 pol2-4 rev3 Delta as compared to pol2-4 rev3 Delta and dpb2-100 rev3 Delta 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 epsilon and GINS is weakened, the access of Pol delta to the leading strand may be increased. The increased participation of Pol delta on the leading strand in the dpb2-100 mutant may explain the synergistic mutator effect observed in the dpb2-100 pol3-5DV double mutant. (C) 2015 The Authors. Published by Elsevier B.V. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).