Both High-Fidelity Replicative and Low-Fidelity Y-Family Polymerases Are Involved in DNA Rereplication

Abstract

DNA rereplication is a major form of aberrant replication that causes genomic instabilities, such as gene amplification. However, little is known about which DNA polymerases are involved in the process. Here, we report that low-fidelity Y-family polymerases (Y-Pols), Pol η, Pol ι, Pol κ, and REV1, significantly contribute to DNA synthesis during rereplication, while the replicative polymerases, Pol δ and Pol ε, play an important role in rereplication, as expected. When rereplication was induced by depletion of geminin, these polymerases were recruited to rereplication sites in human cell lines. This finding was supported by RNA interference (RNAi)-mediated knockdown of the polymerases, which suppressed rereplication induced by geminin depletion. Interestingly, epistatic analysis indicated that Y-Pols collaborate in a common pathway, independently of replicative polymerases. We also provide evidence for a catalytic role for Pol η and the involvement of Pol η and Pol κ in cyclin E-induced rereplication. Collectively, our findings indicate that, unlike normal S-phase replication, rereplication induced by geminin depletion and oncogene activation requires significant contributions of both Y-Pols and replicative polymerases. These findings offer important mechanistic insights into cancer genomic instability.

ACKNOWLEDGMENTS

We thank K. Tomizawa for technical help and H. Ohmori, H. Miyoshi, and B. Vogelstein for generous gifts of materials and reagents.

This work was supported by a Grant-in-Aid for Scientific Research from Japan Science Technology; a grant from the Global COE Program; and a Grant-in-Aid for Scientific Research on Innovative Areas from the Ministry of Education, Science, Technology, Sports and Culture of Japan. Financial support was also provided by the Uehara Memorial Foundation and the Kato Memorial Bioscience Foundation.