title: Impact of DNA3'pp5'G capping on repair reactions at DNA 3' ends.
abstract: Many biological scenarios generate "dirty" DNA 3'-PO4 ends that cannot be sealed by classic DNA ligases or extended by DNA polymerases. The noncanonical ligase RtcB can "cap" these ends via a unique chemical mechanism entailing transfer of GMP from a covalent RtcB-GMP intermediate to a DNA 3'-PO4 to form DNA3'pp5'G. Here, we show that capping protects DNA 3' ends from resection by Escherichia coli exonucleases I and III and from end-healing by T4 polynucleotide 3' phosphatase. By contrast, the cap is an effective primer for DNA synthesis. E. coli DNA polymerase I and Mycobacterium DinB1 extend the DNAppG primer to form an alkali-labile DNApp(rG)pDNA product. The addition of dNTP depends on pairing of the cap guanine with an opposing cytosine in the template strand. Aprataxin, an enzyme implicated in repair of A5'pp5'DNA ends formed during abortive ligation by classic ligases, is highly effective as a DNA 3' decapping enzyme, converting DNAppG to DNA3'p and GMP. We conclude that the biochemical impact of DNA capping is to prevent resection and healing of a 3'-PO4 end, while permitting DNA synthesis, at the price of embedding a ribonucleotide and a pyrophosphate linkage in the repaired strand. Aprataxin affords a means to counter the impact of DNA capping.
authors: Molecular Biology Program, Sloan-Kettering Institute, New York, NY 10065.
authors: Ushati
authors: U
authors: Das
authors: Molecular Biology Program, Sloan-Kettering Institute, New York, NY 10065.
authors: Mathieu
authors: M
authors: Chauleau
authors: Molecular Biology Program, Sloan-Kettering Institute, New York, NY 10065.
authors: Heather
authors: H
authors: Ordonez
authors: Molecular Biology Program, Sloan-Kettering Institute, New York, NY 10065 s-shuman@ski.mskcc.org.
authors: Stewart
authors: S
authors: Shuman
mesh_terms: D001483:Base Sequence; D004247:DNA / Q000096:biosynthesis / Q000378:metabolism*; D004260:DNA Repair; D004259:DNA-Directed DNA Polymerase / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology; D005090:Exodeoxyribonucleases / Q000378:metabolism; D005092:Exonucleases / Q000378:metabolism; D006151:Guanosine / Q000378:metabolism*; D008969:Molecular Sequence Data; D009711:Nucleotides / Q000378:metabolism; D010749:Phosphoprotein Phosphatases / Q000378:metabolism; D012315:RNA Caps / Q000378:metabolism*; D029702:Schizosaccharomyces pombe Proteins / Q000378:metabolism; D013698:Templates, Genetic
chemical_list: D009711:Nucleotides; D012315:RNA Caps; D029702:Schizosaccharomyces pombe Proteins; D006151:Guanosine; D004247:DNA; D004259:DNA-Directed DNA Polymerase; D005090:Exodeoxyribonucleases; D005092:Exonucleases; C569391:Hnt3 protein, S pombe; D010749:Phosphoprotein Phosphatases
keywords: 3′ exonuclease; DNA repair