The rise and fall of poly(ADP-ribose): An enzymatic perspective
Introduction
Cells respond instantaneously to DNA damage with post translational modifications of proteins that repair DNA damage, alter gene expression, or control passage through the cell cycle. The covalent modification of these proteins induce a dynamic network of protein–protein interactions and regulates enzymatic activities, broadly changing cellular physiology and serving to integrate myriad responses to DNA damage that dictate outcomes for DNA repair, cell survival, and responses to chemotherapy. One of the most prodigious posttranslational modifications caused by DNA damage is the poly-(ADP-ribosylation) of proteins, catalyzed by members of the poly-(ADP-ribose) polymerase (PARP) superfamily of NAD+ dependent ADP-ribosyltransferases [1]. Poly-(ADP-ribose) (PAR) is a large, negatively-charged and branched polymer that can exceed the mass of the unmodified protein. PARylation creates binding sites for PAR-specific binding proteins [2], [3] and changes the electrostatic properties of the modified protein, with the notable capacity to change DNA binding properties of enzymes, histones, and structural proteins [4]. PARP-1 itself is the target of most of the poly-(ADP-ribosylation) (PARylation) occurring in response to DNA damage. Automodification of PARP-1 increases its association with a variety of repair and signaling proteins that are recruited to sites of DNA damage by PARP-1 activity [3], [5]. In turn, some of these proteins are PARylated by PARP-1.
PARP enzymes responding to damage can consume substantial amounts of cellular NAD+ within minutes, changing a cell's metabolic status while modifying vast numbers of proteins, many of which have been recently identified by proteomic surveys [6], [7]. For most of these proteins, the effects of PARylation remain to be functionally characterized. These studies are complicated by the fact that PAR modifications turn over rapidly due to the activity of poly-(ADP-ribose) glycohydrolase (PARG) and mono-(ADP-ribose) glycohydrolases (MARGs) [8], [9]. Both the synthesis and turnover of poly-(ADP-ribose) appear to be important for normal responses to DNA damage. In this short perspective, we will review the recent literature on the structures and functions of DNA damage-dependent PARPs and PARG, and then speculate about how these activities may be tied mechanistically to various disease processes and the resulting opportunities for therapeutic intervention.
Section snippets
Structure and mechanism of DNA damage-dependent PARPs
Three members of the PARP superfamily are catalytically activated through interaction with DNA damage: PARP-1, PARP-2, and PARP-3. PARP involvement in the cellular response to DNA damage has long been appreciated and continues to actively develop [10], [11]. A general model that has collectively emerged indicates that the DNA-damage dependent PARPs act early in the process of damage detection, which promptly results in PARP catalytic activation and a burst of PAR production. PARP presence and
Mechanism of PARP-1 activation
Outside of the catalytic domain, the DNA-damage dependent PARPs also have in common a Trp-Gly-Arg (WGR) domain that is essential to damage-dependent activation, and is the most defining feature of the DNA-damage dependent PARPs. A crystal structure that contained the essential domains of PARP-1 in complex with DNA damage provided the first views of the WGR domain contacts with DNA (Fig. 2). The structure indicated that conserved regions of the WGR make sequence-independent contacts with the DNA
Turnover of poly-(ADP-ribose) is required for normal responses to DNA damage
The enzymatic synthesis of poly-(ADP-ribose) and its degradation is commensurately important for normal responses to DNA damage. In mammals, the enzyme poly-(ADP-ribose) glycohydrolase (PARG) is the main activity that removes poly-(ADP-ribose) from proteins by cleaving ribose
ribose bonds [8]. PARG is an abundant enzyme that degrades PAR by a combination of endo- and exo- glycohydrolase activity, removing most of the PAR polymer but leaving a single ADP-ribose attached to the protein. The
Structure and mechanism of PARG
The crystal structure of a bacterial PARG from Thermomonospora curvata [43] revealed an evolutionarily conserved fold that is representative of the core structures of mammalian and Tetrahymena PARG enzymes [44], [45], [46], [47] (Fig. 3A). The catalytic domains of these enzymes share a mixed α, β architecture resembling a Rossman fold, originally termed as macro domain in the transcriptionally repressive histone protein variant, macro-H2A [48]. The macro domain fold binds to ADP-ribose monomers
PAR degradation and DNA repair
During the DNA damage response, PARG activity reverses the automodification of DNA bound PARP-1, concurrent with poly-ubiquitinylation of PARP-1 by the E3 ligase CHFR, and subsequent proteasomal degradation of PARP-1 [56]. Decreased activity of either CHFR or PARG delays repair and causes hypersensitivity to DNA damage [41], [57], [58], [59], indicating that transient PARylation of PARP-1 and the subsequent removal of PARP-1 from DNA strongly contribute to the repair of DNA strand breaks. How
PAR turnover and cell death
PAR oligomers and ADP-ribose are the products of PARG endo- and exo- glycohydrolase activities, respectively. The cellular levels of these metabolites could increase substantially when PARP-1 is hyperactive because PAR is rapidly degraded by PARG [12]. Oligo-PAR has been posited as a signaling molecule that triggers a caspase-independent pathway of programmed cell death, termed necroptosis or parthanatos [50], [51]. Cell death resulting from PARP-1 hyperactivation is typically associated with
Therapeutic interventions directed at poly (ADP-ribose) metabolism
A growing number of PARP inhibitors in clinical trials show promise for the treatment of cancer, although the exact mechanism(s) of their tumor-selective killing effects remain enigmatic [72]. Inhibitors targeting the active site of PARP-1 suffer from dose limiting toxicity, which may result from inadequate binding specificity and off target effects on other PARP family members. As an alternative strategy, inhibitors blocking the DNA-dependent, allosteric activation of PARP-1 enzymatic activity
Acknowledgements
The authors’ work described in this review was funded by grants from the National Institutes of health (R01GM52504 to TE; R01GM087282 to JP; P01-CA092584 to John Tainer, Lawrence Berkeley National Laboratory).
References (75)
- et al.
Reprogramming cellular events by poly(ADP-ribose)-binding proteins
Mol. Aspects Med.
(2013) - et al.
Roles of poly(ADP-ribose) glycohydrolase in DNA damage and apoptosis
Int. Rev. Cell Mol. Biol.
(2013) - et al.
The diverse roles and clinical relevance of PARPs in DNA damage repair: current state of the art
Biochem. Pharmacol.
(2012) Protein ADP-ribosylation and the cellular response to DNA strand breaks
DNA Repair (Amst.)
(2014)- et al.
Structural biology of the writers, readers, and erasers in mono- and poly(ADP-ribose) mediated signaling
Mol. Aspects Med.
(2013) - et al.
PARP-3 and APLF function together to accelerate nonhomologous end-joining
Mol. Cell
(2011) - et al.
PARP-1 mechanism for coupling DNA damage detection to poly(ADP-ribose) synthesis
Curr. Opin. Struct. Biol.
(2013) - et al.
Structural and biophysical studies of human PARP-1 in complex with damaged DNA
J. Mol. Biol.
(2010) - et al.
The DNA-binding domain of human PARP-1 interacts with DNA single-strand breaks as a monomer through its second zinc finger
J. Mol. Biol.
(2011) - et al.
DNA-dependent SUMO modification of PARP-1
DNA Repair (Amst.)
(2013)