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Beyond translesion synthesis: polymerase ? fidelity as a potential determinant of microsatellite stability.

ABSTRACT: Microsatellite DNA synthesis represents a significant component of human genome replication that must occur faithfully. However, yeast replicative DNA polymerases do not possess high fidelity for microsatellite synthesis. We hypothesized that the structural features of Y-family polymerases that facilitate accurate translesion synthesis may promote accurate microsatellite synthesis. We compared human polymerases ? (Pol ?) and ? (Pol ?) fidelities to that of replicative human polymerase ? holoenzyme (Pol ?4), using the in vitro HSV-tk assay. Relative polymerase accuracy for insertion/deletion (indel) errors within 2-3 unit repeats internal to the HSV-tk gene concurred with the literature: Pol ?4 >> Pol ? or Pol ?. In contrast, relative polymerase accuracy for unit-based indel errors within [GT](10) and [TC](11) microsatellites was: Pol ? ? Pol ?4 > Pol ?. The magnitude of difference was greatest between Pols ? and ?4 with the [GT] template. Biochemically, Pol ? displayed less synthesis termination within the [GT] allele than did Pol ?4. In dual polymerase reactions, Pol ? competed with either a stalled or moving Pol ?4, thereby reducing termination. Our results challenge the ideology that pol ? is error prone, and suggest that DNA polymerases with complementary biochemical properties can function cooperatively at repetitive sequences.

SUBMITTER: Hile SE

PROVIDER: S-EPMC3287198 | biostudies-literature | 2012 Feb

REPOSITORIES: biostudies-literature

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Beyond translesion synthesis: polymerase κ fidelity as a potential determinant of microsatellite stability.

Hile Suzanne E SE Wang Xiaoxiao X Lee Marietta Y W T MY Eckert Kristin A KA

Nucleic acids research 20111022 4

Microsatellite DNA synthesis represents a significant component of human genome replication that must occur faithfully. However, yeast replicative DNA polymerases do not possess high fidelity for microsatellite synthesis. We hypothesized that the structural features of Y-family polymerases that facilitate accurate translesion synthesis may promote accurate microsatellite synthesis. We compared human polymerases κ (Pol κ) and η (Pol η) fidelities to that of replicative human polymerase δ holoenzy ...[more]

PMID: 22021378

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Project description:Transcription inhibition by platinum anticancer drugs is an important component of their mechanism of action. Phenanthriplatin, a cisplatin derivative containing phenanthridine in place of one of the chloride ligands, forms highly potent monofunctional adducts on DNA having a structure and spectrum of anticancer activity distinct from those of the parent drug. Understanding the functional consequences of DNA damage by phenanthriplatin for the normal functions of RNA polymerase II (Pol II), the major cellular transcription machinery component, is an important step toward elucidating its mechanism of action. In this study, we present the first systematic mechanistic investigation that addresses how a site-specific phenanthriplatin-DNA d(G) monofunctional adduct affects the Pol II elongation and transcriptional fidelity checkpoint steps. Pol II processing of the phenanthriplatin lesion differs significantly from that of the canonical cisplatin-DNA 1,2-d(GpG) intrastrand cross-link. A majority of Pol II elongation complexes stall after successful addition of CTP opposite the phenanthriplatin-dG adduct in an error-free manner, with specificity for CTP incorporation being essentially the same as for undamaged dG on the template. A small portion of Pol II undergoes slow, error-prone bypass of the phenanthriplatin-dG lesion, which resembles DNA polymerases that similarly switch from high-fidelity replicative DNA processing (error-free) to low-fidelity translesion DNA synthesis (error-prone) at DNA damage sites. These results provide the first insights into how the Pol II transcription machinery processes the most abundant DNA lesion of the monofunctional phenanthriplatin anticancer drug candidate and enrich our general understanding of Pol II transcription fidelity maintenance, lesion bypass, and transcription-derived mutagenesis. Because of the current interest in monofunctional, DNA-damaging metallodrugs, these results are of likely relevance to a broad spectrum of next-generation anticancer agents being developed by the medicinal inorganic chemistry community.

Dataset Information

Beyond translesion synthesis: polymerase ? fidelity as a potential determinant of microsatellite stability.

Publications

Beyond translesion synthesis: polymerase κ fidelity as a potential determinant of microsatellite stability.

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