Differential Activities of DNA Polymerases in Processing Ribonucleotides during DNA Synthesis in Archaea

JOURNAL OF MOLECULAR BIOLOGY

卷:430

Article

Lemor, Melanie¹  Kong, Ziqing²  Henry, Etienne¹  Brizard, Raphael¹  Laurent, Sebastien¹  Bosse, Audrey¹  Henneke, Ghislaine¹ 

[1] Univ Brest, CNRS, Lab Microbiol Environm Extremes, IFREMER, F-29280 Plouzane, France

[2] Umea Univ, Dept Med Biochem & Biophys, Umea, Sweden

关键词: Archaea;    DNA replication and repair;    DNA polymerase;    Nucleotide pool;    Translesion synthesis;   

DOI  :  10.1016/j.jmb.2018.10.004

来源: Elsevier

【 摘 要 】

Consistent with the fact that ribonucleotides (rNTPs) are in excess over deoxyribonucleotides (dNTPs) in vivo, recent findings indicate that replicative DNA polymerases (DNA Pols) are able to insert ribonucleotides (rNMPs) during DNA synthesis, raising crucial questions about the fidelity of DNA replication in both Bacteria and Eukarya. Here, we report that the level of rNTPs is 20-fold higher than that of dNTPs in Pyrococcus abyssi cells. Using dNTP and rNTP concentrations present in vivo, we recorded rNMP incorporation in a template specific manner during in vitro synthesis, with the family-D DNA Pol (PolD) having the highest propensity compared with the family-B DNA Pol and the p41/p46 complex. We also showed that ribonucleotides accumulate at a relatively high frequency in the genome of wild-type Thermococcales cells, and this frequency significantly increases upon deletion of RNase HII, the major enzyme responsible for the removal of RNA from DNA. Because ribonucleotides remain in genomic DNA, we then analyzed the effects on polymerization activities by the three DNA Pols. Depending on the identity of the base and the sequence context, all three DNA Pols bypass rNMP-containing DNA templates with variable efficiency and nucleotide (mis)incorporation ability. Unexpectedly, we found that PoID correctly base-paired a single ribonucleotide opposite rNMP-containing DNA templates. An evolutionary scenario is discussed concerning rNMP incorporation into DNA and genome stability. (C) 2018 The Authors. Published by Elsevier Ltd.

【 授权许可】

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