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Translesion synthesis of abasic sites by yeast DNA polymerase epsilon.

ABSTRACT: Studies of replicative DNA polymerases have led to the generalization that abasic sites are strong blocks to DNA replication. Here we show that yeast replicative DNA polymerase epsilon bypasses a model abasic site with comparable efficiency to Pol eta and Dpo4, two translesion polymerases. DNA polymerase epsilon also exhibited high bypass efficiency with a natural abasic site on the template. Translesion synthesis primarily resulted in deletions. In cases where only a single nucleotide was inserted, dATP was the preferred nucleotide opposite the natural abasic site. In contrast to translesion polymerases, DNA polymerase epsilon with 3'-5' proofreading exonuclease activity bypasses only the model abasic site during processive synthesis and cannot reinitiate DNA synthesis. This characteristic may allow other pathways to rescue leading strand synthesis when stalled at an abasic site.

SUBMITTER: Sabouri N

PROVIDER: S-EPMC2797225 | biostudies-literature | 2009 Nov

REPOSITORIES: biostudies-literature

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Translesion synthesis of abasic sites by yeast DNA polymerase epsilon.

Sabouri Nasim N Johansson Erik E

The Journal of biological chemistry 20090923 46

Studies of replicative DNA polymerases have led to the generalization that abasic sites are strong blocks to DNA replication. Here we show that yeast replicative DNA polymerase epsilon bypasses a model abasic site with comparable efficiency to Pol eta and Dpo4, two translesion polymerases. DNA polymerase epsilon also exhibited high bypass efficiency with a natural abasic site on the template. Translesion synthesis primarily resulted in deletions. In cases where only a single nucleotide was inser ...[more]

PMID: 19776424

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The choice of nucleotide inserted opposite abasic sites formed within chromosomal DNA reveals the polymerase activities participating in translesion DNA synthesis.

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Expression of the cancer-associated DNA polymerase epsilon P286R in fission yeast leads to translesion polymerase dependent hypermutation and defective DNA replication

Project description:Somatic mutations in the proofreading domain of the replicative DNA polymerase epsilon (POLE-exonuclease domain mutations, POLE-EDMs) are frequently found in colorectal and endometrial cancers and, occasionally, in other tumours. POLE-associated cancers typically display hypermutation, microsatellite stability and a unique mutational signature, with predominance of C > A transversions in the context TCT. To understand better the contribution of hypermutagenesis to tumour development, we have modelled the most recurrent POLE-EDM (POLE-P286R) in Schizosaccharomyces pombe. Whole-genome sequencing analysis revealed that the corresponding pol2-P287R also has a strong mutator effect in vivo, with high frequency of base substitutions and relatively few frameshift mutations. The mutations are equally distributed across different genomic regions, but they occur within an AT-rich context. The most abundant mutations are TCT > TAT transversions and, in contrast to human, TCG > TTG transitions are not elevated, likely due to the absence of cytosine methylation in fission yeast. The high mutation burden of pol2-P287R leads to increased sensitivity to elevated dNTP levels and DNA damaging agents and a phenotype that is exacerbated by depletion of the Pfh1 helicase. In addition, S phase is aberrant and RPA foci are elevated, suggestive of persistent ssDNA or DNA damage, and pol2-P287R shows synthetic lethality with rad3 deletion, indicative of checkpoint activation. Significantly, deletion of translesion polymerases kappa and eta partially suppresses pol2-P287R hypermutation, indicating that both enzymes contribute to this phenotype.

2021-06-11 | GSE169231 | GEO

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