ABSTRACT
DNA-dependent poly(ADP-ribose) polymerases (PARPs) PARP1, PARP2 and PARP3 act as DNA break sensors signalling DNA damage. Upon detecting DNA damage, these PARPs use nicotine adenine dinucleotide as a substrate to synthesise a monomer or polymer of ADP-ribose (MAR or PAR, respectively) covalently attached to the acceptor residue of target proteins. Recently, it was demonstrated that PARP1-3 proteins can directly ADP-ribosylate DNA breaks by attaching MAR and PAR moieties to terminal phosphates. Nevertheless, little is still known about the mechanisms governing substrate recognition and specificity of PARP1, which accounts for most of cellular PARylation activity. Here, we characterised PARP1-mediated DNA PARylation of DNA duplexes containing various types of breaks at different positions. The 3'-terminal phosphate residue at double-strand DNA break ends served as a major acceptor site for PARP1-catalysed PARylation depending on the orientation and distance between DNA strand breaks in a single DNA molecule. A preference for ADP-ribosylation of DNA molecules containing 3'-terminal phosphate over PARP1 auto-ADP-ribosylation was observed, and a model of DNA modification by PARP1 was proposed. Similar results were obtained with purified recombinant PARP1 and HeLa cell-free extracts. Thus, the biological effects of PARP-mediated ADP-ribosylation may strongly depend on the configuration of complex DNA strand breaks.
Subject(s)
DNA/chemistry , Poly (ADP-Ribose) Polymerase-1/chemistry , Animals , Catalysis , Cattle , Humans , Substrate SpecificityABSTRACT
Covalent linkage of ADP-ribose units to proteins catalyzed by poly(ADP-ribose) polymerases (PARPs) plays important signaling functions in a plethora of cellular processes including DNA damage response, chromatin organization, and gene transcription. Poly- and mono-ADP-ribosylation of target macromolecules are often responsible both for the initiation and for coordination of these processes in mammalian cells. Currently, the number of cellular targets for ADP-ribosylation is rapidly expanding, and the molecular mechanisms underlying the broad substrate specificity of PARPs present enormous interest. In this review, the roles of PARP-mediated modifications of protein and nucleic acids, the readers of ADP-ribosylated structures, and the origin and function of programmed DNA strand breaks in PARP activation, transcription regulation, and DNA demethylation are discussed.
