Spatial and functional relationship between poly(ADP-ribose) polymerase-1 and poly(ADP-ribose) glycohydrolase in the brain

Abstract

Poly(ADP-ribose) polymerases (PARPs) are members of a family of enzymes that utilize NAD + as substrate to form large ADP-ribose polymers (PAR) in the nucleus. PAR has a very short half life due to its rapid degradation by poly(ADP-ribose) glycohydrolase (PARG). PARP-1 mediates acute neuronal cell death induced by a variety of insults including cerebral ischemia, MPTP-induced Parkinsonism, and CNS trauma. While PARP-1 is localized to the nucleus, PARG resides in both the nucleus and cytoplasm. Surprisingly, there appears to be only one gene encoding PARG activity, which has been characterized in vitro to generate different splice variants, in contrast to the growing family of PARPs. Little is known regarding the spatial and functional relationships of PARG and PARP-1. Here we evaluate PARG expression in the brain and its cellular and subcellular distribution in relation to PARP-1. Anti-PARG (α-PARG) antibodies raised in rabbits using a purified 30 kDa C-terminal fragment of murine PARG recognize a single band at 111 kDa in the brain. Western blot analysis also shows that PARG and PARP-1 are evenly distributed throughout the brain. Immunohistochemical studies using α-PARG antibodies reveal punctate cytosolic staining, whereas anti-PARP-1 (α-PARP-1) antibodies demonstrate nuclear staining. PARG is enriched in the mitochondrial fraction together with manganese superoxide dismutase (MnSOD) and cytochrome C (Cyt C) following whole brain subcellular fractionation and Western blot analysis. Confocal microscopy confirms the co-localization of PARG and Cyt C. Finally, PARG translocation to the nucleus is triggered by NMDA-induced PARP-1 activation. Therefore, the subcellular segregation of PARG in the mitochondria and PARP-1 in the nucleus suggests that PARG translocation is necessary for their functional interaction. This translocation is PARP-1 dependent, further demonstrating a functional interaction of PARP-1 and PARG in the brain.

PARP and PARG Inhibitors—New Therapeutic Targets in Cancer Treatment

“…To date, only one single PARG gene has been detected in mammals [5] encoding for 3 cDNAs which generates 3 isoforms namely 110 kDa, 102 kDa and 99 kDa [6], amongst which its major form is 110 kDa. The latter has been localized to the nucleus, the cytoplasm [7]; co-localised with cytochrome C [8] and during mitosis has been identified in the spindle body and centrosomes too [9]. However, human PARG known as full length 111 kDa is confined to the nucleus due to a NLS (nuclear localization signal) encoded by exon1 [10].…”

“…However, human PARG known as full length 111 kDa is confined to the nucleus due to a NLS (nuclear localization signal) encoded by exon1 [10]. It has been found that PARG is not only expressed in different subcellular compartments of the neuronal tissues but also heart, testis and kidneys; mainly in their mitochondrial/cytosolic fraction [8]. Since, the poly ADPribosylation of nuclear proteins is an ongoing process; so, nuclear-cytosolic shuttling of PARG is constant and at the same time, the endo-glycosylase activity of PARG which is physiologically important continuously generates protein free ADPr polymers that can interact with histones and other nuclear proteins useful in intracellular pathways [1].…”

Mitochondrial Localization of PARP-1 Requires Interaction with Mitofilin and Is Involved in the Maintenance of Mitochondrial DNA Integrity

“…However, despite the clear involvement of PAR signaling in mitochondrial dysfunction, it is still debated whether these effects are mediated by a mitochondrially localized PARP-1. In fact, although some studies provided evidence for the presence of PARP-1 in mitochondria (18,20,21), others failed to detect the enzyme in this organelle, thus suggesting that the mitochondrial effects of PARP-1 activation are triggered by the nuclear enzyme (6,22,23).…”

Functional Localization of Two Poly(ADP-Ribose)-Degrading Enzymes to the Mitochondrial Matrix

“…According to immunocytochemical studies, at least one isoform was clearly associated with mitochondria, although the suborganellar localization, and consequently its function, has remained unknown (33). A recent report suggested the dissociation of a PARG isoform from mitochondria following PARP1 activation in neuronal cells (44).…”