Abstract
Poly(ADP-ribose) polymerase-1 (PARP-1) activation is a hallmark of oxidative stress-induced cellular injury that can lead to energetic failure and necrotic cell death via depleting the cellular nicotinamide adenine dinucleotide (NAD(+)) and ATP pools. Pharmacological PARP-1 inhibition or genetic PARP-1 deficiency exert protective effects in multiple models of cardiomyocyte injury. However, the connection between nuclear PARP-1 activation and depletion of the cytoplasmic and mitochondrial energy pools is poorly understood. By using cultured rat cardiomyocytes, here we report that ring finger protein 146 (RNF146), a cytoplasmic E3-ubiquitin ligase, acts as a direct interactor of PARP-1. Overexpression of RNF146 exerts protection against oxidant-induced cell death, whereas PARP-1-mediated cellular injury is augmented after RNF146 silencing. RNF146 translocates to the nucleus upon PARP-1 activation, triggering the exit of PARP-1 from the nucleus, followed by rapid degradation of both proteins. PARP-1 and RNF146 degradation occurs in the early phase of myocardial ischemia-reperfusion injury; it precedes the induction of heat shock protein expression. Taken together, PARP-1 release from the nucleus and its rapid degradation represent newly identified steps of the necrotic cell death program induced by oxidative stress. These steps are controlled by the ubiquitin-proteasome pathway protein RNF146. The current results shed new light on the mechanism of necrotic cell death. RNF146 may represent a distinct target for experimental therapeutic intervention of oxidant-mediated cardiac injury.
Highlights
Poly(ADP-ribose) polymerase-1 (PARP-1) is a ubiquitously expressed enzyme that catalyzes the poly(ADPribosyl)ation of acceptor proteins by using nicotinamide adenine dinucleotide (NAD+) as a substrate
Because our pilot studies showed that ring finger protein 146 (RNF146) expression is the highest in the heart and muscle, the goal of the current project was to characterize the role of RNF146 in PARP-1–mediated cell death during oxidative myocyte injury in vitro and during myocardial ischemia-reperfusion injury in vivo
Because RNF146 has previously been demonstrated to be protective against PARP-1–mediated cell death in neurons, we tested whether it exerts similar protection against oxidant injury in cardiomyocytes by using siRNA-mediated gene silencing and stable transfection of H9c2 cells with RNF146 expression vector (Figure 2)
Summary
Poly(ADP-ribose) polymerase-1 (PARP-1) is a ubiquitously expressed enzyme that catalyzes the poly(ADPribosyl)ation of acceptor proteins by using nicotinamide adenine dinucleotide (NAD+) as a substrate. The protein consists of an N-terminal DNA-binding domain, an automodification domain and a C-terminal catalytic domain. PARP-1 has low basal enzymatic activity, but its catalytic activity is dramatically stimulated on binding to damaged DNA (single or double strand breaks). Targets of the enzyme include histone proteins and transcription-related factors and PARP-1 itself (via its automodification domain). PARylation can affect the target protein function and its interactions with various proteins and DNA; thereby, PARP-1 plays a key role in the regulation of DNA repair and gene transcription [1,2]. The regulation of nuclear DNA repair and maintenance of genomic integrity was considered the main physiological function of PARP-1.
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