Dataset Information

Selective down-regulation of nuclear poly(ADP-ribose) glycohydrolase.

ABSTRACT: The formation of ADP-ribose polymers on target proteins by poly(ADP-ribose) polymerases serves a variety of cell signaling functions. In addition, extensive activation of poly(ADP-ribose) polymerase-1 (PARP-1) is a dominant cause of cell death in ischemia-reperfusion, trauma, and other conditions. Poly(ADP-ribose) glycohydrolase (PARG) degrades the ADP-ribose polymers formed on acceptor proteins by PARP-1 and other PARP family members. PARG exists as multiple isoforms with differing subcellular localizations, but the functional significance of these isoforms is uncertain.Primary mouse astrocytes were treated with an antisense phosphorodiamidate morpholino oligonucleotide (PMO) targeted to exon 1 of full-length PARG to suppress expression of this nuclear-specific PARG isoform. The antisense-treated cells showed down-regulation of both nuclear PARG immunoreactivity and nuclear PARG enzymatic activity, without significant alteration in cytoplasmic PARG activity. When treated with the genotoxic agent MNNG to induced PARP-1 activation, the antisense-treated cells showed a delayed rate of nuclear PAR degradation, reduced nuclear condensation, and reduced cell death.These results support a preferentially nuclear localization for full-length PARG, and suggest a key role for this isoform in the PARP-1 cell death pathway.

SUBMITTER: Burns DM

PROVIDER: S-EPMC2655720 | biostudies-literature | 2009

REPOSITORIES: biostudies-literature

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Selective down-regulation of nuclear poly(ADP-ribose) glycohydrolase.

Burns David M DM Ying Weihai W Kauppinen Tiina M TM Zhu Keqing K Swanson Raymond A RA

PloS one 20090325 3

<h4>Background</h4>The formation of ADP-ribose polymers on target proteins by poly(ADP-ribose) polymerases serves a variety of cell signaling functions. In addition, extensive activation of poly(ADP-ribose) polymerase-1 (PARP-1) is a dominant cause of cell death in ischemia-reperfusion, trauma, and other conditions. Poly(ADP-ribose) glycohydrolase (PARG) degrades the ADP-ribose polymers formed on acceptor proteins by PARP-1 and other PARP family members. PARG exists as multiple isoforms with dif ...[more]

PMID: 19319190

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Project description:BackgroundInitiation, amplitude, duration and termination of transforming growth factor ? (TGF?) signaling via Smad proteins is regulated by post-translational modifications, including phosphorylation, ubiquitination and acetylation. We previously reported that ADP-ribosylation of Smads by poly(ADP-ribose) polymerase 1 (PARP-1) negatively influences Smad-mediated transcription. PARP-1 is known to functionally interact with PARP-2 in the nucleus and the enzyme poly(ADP-ribose) glycohydrolase (PARG) can remove poly(ADP-ribose) chains from target proteins. Here we aimed at analyzing possible cooperation between PARP-1, PARP-2 and PARG in regulation of TGF? signaling.MethodsA robust cell model of TGF? signaling, i.e. human HaCaT keratinocytes, was used. Endogenous Smad3 ADP-ribosylation and protein complexes between Smads and PARPs were studied using proximity ligation assays and co-immunoprecipitation assays, which were complemented by in vitro ADP-ribosylation assays using recombinant proteins. Real-time RT-PCR analysis of mRNA levels and promoter-reporter assays provided quantitative analysis of gene expression in response to TGF? stimulation and after genetic perturbations of PARP-1/-2 and PARG based on RNA interference.ResultsTGF? signaling rapidly induces nuclear ADP-ribosylation of Smad3 that coincides with a relative enhancement of nuclear complexes of Smads with PARP-1 and PARP-2. Inversely, PARG interacts with Smads and can de-ADP-ribosylate Smad3 in vitro. PARP-1 and PARP-2 also form complexes with each other, and Smads interact and activate auto-ADP-ribosylation of both PARP-1 and PARP-2. PARP-2, similar to PARP-1, negatively regulates specific TGF? target genes (fibronectin, Smad7) and Smad transcriptional responses, and PARG positively regulates these genes. Accordingly, inhibition of TGF?-mediated transcription caused by silencing endogenous PARG expression could be relieved after simultaneous depletion of PARP-1.ConclusionNuclear Smad function is negatively regulated by PARP-1 that is assisted by PARP-2 and positively regulated by PARG during the course of TGF? signaling.

Dataset Information

Selective down-regulation of nuclear poly(ADP-ribose) glycohydrolase.

Publications

Selective down-regulation of nuclear poly(ADP-ribose) glycohydrolase.

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