ADP‐Ribosylation of Highly Purified Rat Brain Mitochondria

Abstract

Highly purified synaptic and nonsynaptic mitochondria were prepared from rat brain, and their ADP-ribosyl transferase and NAD glycohydrolase activities were investigated. Data show that there is no significant difference in ADP-ribosyl transferase activity between these two types of subcellular preparations. However, NAD glycohydrolase activity appeared to be much higher in nonsynaptic mitochondria. The specific activity of both enzymes was investigated in the presence of the inhibitor nicotinamide or its analogue 3-aminobenzamide or other adenine nucleotides, such as ATP or ADP-ribose. The inhibitory effect of nicotinamide or 3-aminobenzamide on ADP-ribosyl transferase appears rather weak compared with their effect on NAD glycohydrolase activity. However, ADP-ribose and ATP appeared more effective in inhibiting ADP-ribosyl transferase. Our results provide evidence for the existence of ADP-ribosyl transferase activity in rat brain mitochondria. When NAD glycohydrolase was inhibited totally by nicotinamide, the transfer of ADP-ribose from NAD to mitochondrial proteins still occurred. The chain length determinations show that the linkage of ADP-ribose to mitochondrial proteins is oligomeric.

Regulation of Mitochondrial Poly(ADP-Ribose) Polymerase Activation by the β-Adrenoceptor/cAMP/Protein Kinase A Axis during Oxidative Stress

“…Although PARP1 activation and the associated cell death response were initially attributed to events in the nuclear compartment, subsequent studies demonstrated the mitochondrial presence of PARP1 and an independent role for mitochondrial PARP1 in mediating cell death. (Masmoudi et al, 1988;Du et al, 2003;Rossi et al, 2009). Based on the results of the PLA assays, we conclude that the mitochondrial PARP1 activation is an early event in oxidant-induced cell injury and, in line with the extranuclear/mitochondrial localization of PKA (Gancedo, 2013;Taylor et al, 2013;Valsecchi et al, 2013), the PKA/PARP1 interactions primarily occur in the extranuclear/mitochondrial compartment.…”

“…Its pathophysiologic overactivation, as a result of oxidative DNA injury, has been implicated in the pathogenesis of several diseases, including neuroinjury, inflammation, and ischemia/reperfusion, and various forms of critical illness (Tulin et al, 2003;Jagtap and Szabo, 2005;DeVos et al, 2012;Szanto et al, 2012;Curtin and Szabo, 2013;Burkle and Virag, 2013). In addition to the nuclear isoform of PARP1 several sets of studies have identified PARP1 in the mitochondrial compartment as well; it appears that, in addition to nuclear PARP1, mitochondrial PARP1 also plays roles in the regulation of various cellular functions-at least in some cell types (Masmoudi et al, 1988;Du et al, 2003;Rossi et al, 2009, reviewed in Burkle andVirag, 2013).…”

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

“…Furthermore, increasing evidence indicates that mitochondrial DNA, which is constantly exposed to oxidative damage, is efficiently repaired through at least a subset of the mechanisms involved in nuclear DNA repair (14). A number of previous works reported the occurrence of intramitochondrial poly(ADP-ribosyl)ation (15)(16)(17)(18). Moreover, it has been recently established that PARP-1 hyperactivation can cause the release of apoptosis-inducing factor (AIF) from mitochondria, triggering a caspase-independent cell death pathway (19).…”

Intra-mitochondrial Poly(ADP-ribosylation) Contributes to NAD+ Depletion and Cell Death Induced by Oxidative Stress

“…However, the capacity for PARP-1 activation within the nucleus to deplete total cellular energy stores, particularly compartmentalized within mitochondria, remains to be established (4,6). Because in addition to being abundant in cell nuclei, PARP-1 and other ADP-ribosyltransferases are also prevalent in mitochondria (7)(8)(9), where similar to nuclear PARP-1, they facilitate DNA repair in response to oxidative damage (10,11), we hypothesized that inhibition of mitochondrial poly(ADP-ribosylation) may play a pivotal role in neuronal cell survival under conditions of oxidative stress and excitotoxicity.…”