Poly(ADP-ribose) polymerases covalently modify strand break termini in DNA fragments in vitro

Poly(ADP-ribose) polymerases (PARPs/ARTDs) use nicotinamide adenine dinucleotide (NAD ) to catalyse the synthesis of a long branched poly(ADP-ribose) polymer (PAR) attached to the acceptor amino acid residues of nuclear proteins. PARPs act on single- and double-stranded DNA breaks by recruiting DNA...

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Bibliographic Details
Published in:Nucleic acids research 2016-11, Vol.44 (19), p.9279-9295
Main Authors: Talhaoui, Ibtissam, Lebedeva, Natalia A, Zarkovic, Gabriella, Saint-Pierre, Christine, Kutuzov, Mikhail M, Sukhanova, Maria V, Matkarimov, Bakhyt T, Gasparutto, Didier, Saparbaev, Murat K, Lavrik, Olga I, Ishchenko, Alexander A
Format: Article
Language:English
Subjects:
Animals
Biochemistry, Molecular Biology
Catalysis
DNA - genetics
DNA - metabolism
DNA Adducts
DNA Breaks, Double-Stranded
Genome Integrity, Repair and
Genomics
Humans
Hydrolysis
Life Sciences
Mice
NAD - metabolism
Poly (ADP-Ribose) Polymerase-1 - metabolism
Poly(ADP-ribose) Polymerases - metabolism
Protein Binding
Substrate Specificity
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Summary:Poly(ADP-ribose) polymerases (PARPs/ARTDs) use nicotinamide adenine dinucleotide (NAD ) to catalyse the synthesis of a long branched poly(ADP-ribose) polymer (PAR) attached to the acceptor amino acid residues of nuclear proteins. PARPs act on single- and double-stranded DNA breaks by recruiting DNA repair factors. Here, in in vitro biochemical experiments, we found that the mammalian PARP1 and PARP2 proteins can directly ADP-ribosylate the termini of DNA oligonucleotides. PARP1 preferentially catalysed covalent attachment of ADP-ribose units to the ends of recessed DNA duplexes containing 3'-cordycepin, 5'- and 3'-phosphate and also to 5'-phosphate of a single-stranded oligonucleotide. PARP2 preferentially ADP-ribosylated the nicked/gapped DNA duplexes containing 5'-phosphate at the double-stranded termini. PAR glycohydrolase (PARG) restored native DNA structure by hydrolysing PAR-DNA adducts generated by PARP1 and PARP2. Biochemical and mass spectrometry analyses of the adducts suggested that PARPs utilise DNA termini as an alternative to 2'-hydroxyl of ADP-ribose and protein acceptor residues to catalyse PAR chain initiation either via the 2',1″-O-glycosidic ribose-ribose bond or via phosphodiester bond formation between C1' of ADP-ribose and the phosphate of a terminal deoxyribonucleotide. This new type of post-replicative modification of DNA provides novel insights into the molecular mechanisms underlying biological phenomena of ADP-ribosylation mediated by PARPs.
ISSN:0305-1048
1362-4962
DOI:10.1093/nar/gkw675