Analysis of CPD Ultraviolet Lesion Bypass in Chicken DT40 Cells: Polymerase eta and PCNA Ubiquitylation Play Identical Roles

作者：Varga Agnes; Marcus Adam P; Himoto Masayuki; Iwai Shigenori; Szuets David

来源：PLos One, 2012, 7(12): e52472.

DOI：10.1371/journal.pone.0052472

Translesion synthesis (TLS) provides a mechanism of copying damaged templates during DNA replication. This potentially mutagenic process may operate either at the replication fork or at post-replicative gaps. We used the example of T-T cyclobutane pyrimidine dimer (CPD) bypass to determine the influence of polymerase recruitment via PCNA ubiquitylation versus the REV1 protein on the efficiency and mutagenic outcome of TLS. Using mutant chicken DT40 cell lines we show that, on this numerically most important UV lesion, defects in polymerase eta or in PCNA ubiquitylation similarly result in the long-term failure of lesion bypass with persistent strand gaps opposite the lesion, and the elevation of mutations amongst successful TLS events. Our data suggest that PCNA ubiquitylation promotes CPD bypass mainly by recruiting polymerase eta, resulting in the majority of CPD lesions bypassed in an error-free manner. In contrast, we find that polymerase zeta is responsible for the majority of CPD-dependent mutations, but has no essential function in the completion of bypass. These findings point to a hierarchy of access of the different TLS polymerases to the lesion, suggesting a temporal order of their recruitment. The similarity of REV1 and REV3 mutant phenotypes confirms that the involvement of polymerase zeta in TLS is largely determined by its recruitment to DNA by REV1. Our data demonstrate the influence of the TLS polymerase recruitment mechanism on the success and accuracy of bypass.

• 出版日期2012-12-18