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

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Talhaoui I.
Lebedeva N.A.
Zarkovic G.
Saint-Pierre C.
Kutuzov M.M.
Sukhanova M.V.
Matkarimov B.T.
Gasparutto D.
Saparbaev M.K.
Lavrik O.I.
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Oxford University Press
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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.
Fondation ARC ( [PJA20151203415 to A.A.I.]; ERA.Net RUS Plus ( [#306 to A.A.I. and RFBR-16-54-76010 to O.I.L.]; Electricité de France ( [RB 2016-17 to M.K.S.]; Science Committee of the Ministry of Education and Science of the Republic of Kazakhstan (program 0212/PTF-14-OT) [3755/GF4 and 2835/GF3] ( to B.T.M.; RSF [14-24-00038 to O.I.L.]; RFBR [15-54-16003]; Program of RAS on Molecular and Cellular Biology [6.4]; postdoctoral and doctoral fellowships from Fondation ARC ( [PDF20110603195 to I.T. and G.Z.] and CIENCIACTIVA/CONCYTEC (, respectively. Funding for open access charge: National Laboratory Astana, Nazarbayev University, Astana, Republic of Kazakhstan.
Palabras clave
monomer, cordycepin, DNA fragment, double stranded DNA, glycosidase