Dataset Information

Poly(ADP-ribose) polymerase 1 is indispensable for transforming growth factor-? Induced Smad3 activation in vascular smooth muscle cell.

ABSTRACT:

Background

Transforming growth factor type-? (TGF-?)/Smad pathway plays an essential role in vascular fibrosis. Reactive oxygen species (ROS) generation also mediates TGF-? signaling-induced vascular fibrosis, suggesting that some sort of interaction exists between Smad and redox pathways. However, the underlying molecular mechanism is largely unknown. This study aims to investigate the influence of poly(ADP-ribose) polymerase 1 (PARP1), a downstream effector of ROS, on TGF-? signaling transduction through Smad3 pathway in rat vascular smooth muscle cells (VSMCs).

Methods and results

TGF-?1 treatment promoted PARP1 activation through induction of ROS generation in rat VSMCs. TGF-?1-induced phosphorylation and nuclear accumulation of Smad3 was prevented by treatment of cells with PARP inhibitor, 3-aminobenzamide (3AB) or N-(6-oxo-5,6-dihydrophenanthridin-2-yl)-2-(N,N-dimethylamino)acetami (PJ34), or PARP1 siRNA. TGF-?1 treatment promoted poly(ADP-ribosy)lation of Smad3 via activation of PARP1 in the nucleus. Poly(ADP-ribosy)lation enhanced Smad-Smad binding element (SBE) complex formation in nuclear extracts and increased DNA binding activity of Smad3. Pretreatment with 3AB, PJ34, or PARP1 siRNA prevented TGF-?1-induced Smad3 transactivation and expression of Smad3 target genes, including collagen I?1, collagen III?1 and tissue inhibitor of metalloproteinase 1, in rat VSMCs.

Conclusions

PARP1 is indispensable for TGF-?1 induced Smad3 activation in rat VSMCs. Targeting PARP1 may be a promising therapeutic approach against vascular diseases induced by dysregulation of TGF-?/Smad3 pathway.

SUBMITTER: Huang D

PROVIDER: S-EPMC3205050 | biostudies-literature | 2011

REPOSITORIES: biostudies-literature

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Publications

Poly(ADP-ribose) polymerase 1 is indispensable for transforming growth factor-β Induced Smad3 activation in vascular smooth muscle cell.

Huang Dan D Wang Yan Y Wang Lin L Zhang Fengxiao F Deng Shan S Wang Rui R Zhang Yun Y Huang Kai K

PloS one 20111031 10

<h4>Background</h4>Transforming growth factor type-β (TGF-β)/Smad pathway plays an essential role in vascular fibrosis. Reactive oxygen species (ROS) generation also mediates TGF-β signaling-induced vascular fibrosis, suggesting that some sort of interaction exists between Smad and redox pathways. However, the underlying molecular mechanism is largely unknown. This study aims to investigate the influence of poly(ADP-ribose) polymerase 1 (PARP1), a downstream effector of ROS, on TGF-β signaling t ...[more]

PMID: 22073128

Similar Datasets

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.