Search

The anticancer drug cisplatin forms 1,2-d(GpG) DNA intrastrand cross-links (cisplatin lesions) that stall RNA polymerase II (Pol II) and trigger transcription-coupled DNA repair. Here we present a structure-function analysis of Pol II stalling at a cisplatin lesion in the DNA template. Pol II stalling results from a translocation barrier that prevents delivery of the lesion to the active site. AMP misincorporation occurs at the barrier and also at an abasic site, suggesting that it arises from nontemplated synthesis according to an 'A-rule' known for DNA polymerases. Pol II can bypass a cisplatin lesion that is artificially placed beyond the translocation barrier, even in the presence of a G·A mismatch. Thus, the barrier prevents transcriptional mutagenesis. The stalling mechanism differs from that of Pol II stalling at a photolesion, which involves delivery of the lesion to the active site and lesion-templated misincorporation that blocks transcription. [ABSTRACT FROM AUTHOR]

DNA polymerase η (Pot η) is a eukaryotic lesion bypass polymerase that helps organisms to survive exposure to ultraviolet (UV) radiation, and tumor cells to gain resistance against dsplatin-based chemotherapy. It allows cells to replicate across cross-link lesions such as 1,2-d(GpG) cisptatin adducts (Pt-GG) and UV-induced ds-syn thymine dimers. We present structural and biochemical analysis of how Pot η copies Pt-GG-containing DNA. The damaged DNA is bound in an open DNA binding rim. Nucleotidyl transfer requires the DNA to rotate into an active conformation, driven by hydrogen bonding of the temptating base to the dNTP. For the 3′dG of the Pt-GG, this step is accomplished by a Watson-Crick base pair to dCTP and is biochemically efficient and accurate. In contrast, bypass of the 5′dG of the Pt-GG is less efficient and promiscuous for dCTP and dATP as a result of the presence of the rigid Pt cross-link. Our analysis reveals the set of structural features that enable Pot η to replicate across strongly distorting DNA lesions. [ABSTRACT FROM AUTHOR]