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Studies over many years have revealed the central importance of poly(ADP-ribose) metabolism in the maintenance of genomic integrity. While the involvement of poly(ADP-ribose) polymerase-1 (PARP-1) in this metabolism has been long known, more recent studies have demonstrated the contribution of many different genes coding for PARPs to promoting cellular recovery from genotoxic stress, eliminating badly damaged cells from the organism, and ensuring accurate transmission of genetic information during cell division. Additionally, emerging information suggests the involvement of ADP-ribose polymer metabolism in the regulation of intracellular trafficking, memory formation and other cellular functions. This chapter reviews the chemistry of ADP-ribose polymer metabolism and the enzymes that catalyze the synthesis and turnover of poly(ADP-ribose).

Poly(ADP-ribosyl)ation is an immediate DNA damage-dependent posttranslational modification of histones and other nuclear proteins that contributes to the survival of injured proliferating cells. Poly(ADP-ribose) polymerases (PARPs) now constitute a large family of 18 proteins, encoded by different genes and displaying a conserved catalytic domain in which PARP-1 (113 kDa), the founding member, and PARP-2 (62 kDa) are so far the sole enzymes whose catalytic activity is immediately stimulated by DNA strand-breaks. This review summarizes our present knowledge of the structure and function of PARP-2, the closest relative to PARP-1.

Poly(ADP-ribosyl)ation is a post-translational modification of proteins. Upon DNA damage, poly(ADP-ribose) polymerase-1 catalyzes the transfer of ADP-ribose moieties from NAD^+ onto acceptor proteins to form long and branched polymers. Poly(ADP-ribosyl)ation is an extensive but transient modification as polymer chains can reach more than 200 units on protein acceptors and be degraded within a few minutes by poly(ADP-ribose) glycohydrolase. Homeostasis of poly(ADP-ribose) is thought to play an important function in cellular processes. The importance of pADPR synthesis has been established in vitro and in vivo by using chemical inhibitors and genetically engineered mutant mice devoid of the main pADPR synthesizing enzyme, PARP-1. However, the function of PARG in vivo remains elusive. This chapter describes the generation and characterization of PARG knockout mice.

Several lines of evidence reveal that poly(ADP-ribose)polymerase-1 (PARP-1) operates in a DNA damage signaling network. Poly(ADP-ribose) metabolism induced by DNA damage participates in DNA repair and contributes to downstream mechanisms leading to cell cycle arrest, cell survival, cell death, or cell transformation. An important element of these multiple actions is the recruitment of DNA damage checkpoint proteins coordinating DNA repair with downstream events. The focus of this overview is the mechanism by which poly(ADP-ribose)—attached to the automodified PARP-1—interact with DNA damage check-point proteins and how it may reprogram the functions of specific protein domains. Several proteins of the genome surveillance system, e.g., p53, p21, DNA-PK, NF-κB, XRCC1, and XPA are targets of such regulation. In all cass studied, a specific ‘polymer-binding’ sequence motif of 20 to 26 amino acids is targeted by poly (ADP-ribose) and this motif overlaps with important functional domains responsible for protein-protein or protein-DNA interactions, nuclear import or export, enzymatic catalysis, or protein degradation.

The chemical structure of poly(ADP-ribose) suggests not only that its modification of acceptor proteins should modify the structure and function of the acceptor proteins, but also that the poly(ADP-ribose) molecule itself should possess an intrinsic structural information that can alter cellular function(s). The localization of PARP-1 to the centrosome clearly shows that its function is not only confined to the nucleus, but plays a role also in the cytoplasm. Thus poly(ADP-ribosyl)ation should be considered an important regulatory mechanism not only in the nucleus, but in the cell at large. In this context, the interaction between nuclear and cytoplasmic events through the poly(ADP-ribosyl)ation reaction is an intriguing possibility. Understanding poly(ADP-ribose) metabolism has an important impact for unraveling fundamental biological mechanisms ranging from chromosomal instability in cancer, the morphogenesis of the tissues and the maintenance of neuronal cell functions.

Poly(ADP-ribose) polymerase (PARP-1) plays a crucial role in DNA repair and interacts with many DNA replication/repair factors, including the proliferating cell nuclear antigen (PCNA), a protein involved in many DNA transactions. The association between these proteins in vitro results in the inhibition of PARP-1 activity and of PCNA-dependent DNA synthesis carried out by DNA polymerase δ. In vivo, the interaction of the two proteins has been shown to increase after DNA damage. The activity of PCNA is regulated by p21^waf1/cip1, which binds the interdomain connector loop of PCNA in the same region involved in the interaction of PCNA with DNA polymerase δ. This feature enables p21 to compete with DNA polymerase δ to regulate the differential inhibition of DNA replication vs. DNA repair. The finding that PARP- 1 is associated with both PCNA and p21 suggests a possible cooperation of PARP-1 and p21 in regulating the functions of PCNA during DNA replication/repair.

Mammalian poly (ADP-ribose) polymerase-1 (PARP-1) is a nuclear chromatin associated protein and belongs to a large family of enzymes that can synthesize large branched polymers of ADP-ribose units by using β-nicotinamide adenine dinucleotide (NAD^+) as substrate. A pathophysiological role for PARP-1 has been demonstrated in a number of diseases and animal models using PARP-1 knockout mice. The transcription factor nuclear factor kappa B (NF-κB) encompasses a family of inducible transcription factors, which are required for regulation of genes, and has traditionally been linked to cellular immune and inflammatory response. It has been clearly demonstrated that excessive activation or inappropriate regulation of immune and inflammation cascades causes tissue and cellular damage, which may lead to cellular dysfunction and cell death. Recent reports provided strong evidence that PARP-1 is required for NF-κB-dependent gene expression in a stimuli dependent manner and acts as a transcriptional coactivator of NF-κB. These findings might provide new insights in the pathophysiology of different inflammatory disorders.

In the post-genome era attention is being focused on those epigenetic modifications which modulate chromatin structure to guarantee that information present on DNA is read correctly and at the most appropriate time in order to meet cellular requirements. In this chapter data are reviewed which show that along the chain of events that induce DNA methylation-dependent chromatin condensation, a post-synthetic modification other than histone acetylation, phosphorylation and methylation, namely poly(ADP-ribosyl)ation, participates in the establishment and maintenance of methylation-free regions of chromatin. In fact, several lines of in vitro and in vivo evidence have shown that poly(ADP-ribosyl)ation is involved in the control of DNA methylation pattern, protecting genomic DNA from full methylation. Molecular mechanism(s) that might underpin the correlation between inhibition of poly(ADP-ribose) polymerases and DNA hypermethylation will be discussed. Finally the hypothesis is posited that inhibition of the poly(ADP-ribosyl)ation process in the cell may be responsible for the anomalous hypermethylation of tumor suppressor gene promoters during tumorigenesis.

Mitochondria and the nucleus play critical roles in cell death. The signaling between these two organelles is dynamic. Understanding the signaling process between the nucleus and the mitochondria in cell death programs could lead us to more options in the development of new therapeutic targets. We discussed in this chapter how AIF could be released from the mitochondria and how AIF could act in PARP-dependent cell death mechanism. The molecular mechanisms accounting for PARP-dependent cell death are still being investigated but mitochondrial release of AIF and its translocation to the nucleus appears to be essential in this death process. We speculate that the PARP-mediated depletion of NAD and ATP or novel signals from PARP over activation might trigger AIF translocation; however, the authentic factors and the legitimate mechanism of mitochondrial AIF release remain to be explored.