Sirt1 interaction with active Smad2 modulates transforming growth factor-β regulated transcription

Abstract

BackgroundThe simplicity of Transforming Growth Factor ß (TGFβ) signaling pathway, linear and non-amplified, hardly sustains its variety of responses. This is often justified by the complex regulation showed by Smad proteins, TGFβ signaling intracellular transducers, object of post-translational modifications that modulate TGFβ-dependent transcription. Protein acetylation is emerging as a compelling mechanism affecting the activities of significant transcription factors, including p53, FOXO or NF-kB. Smad proteins might be controlled by this mechanism, implying that accessory factors capable of altering Smads-transcriptional complexes acetylation status and hence regulate TGFβ responses remain to be identified. Understanding this interaction may help in the assessment of TGFβ signaling outcomes, extending from healthy physiology to pathological conditions and cancer.MethodsA two-hybrid chimera interacting system allowed to identify Sirt1, a NAD+ dependent type III histone deacetylase, as a novel Smad2 interactor. Several well stablished cellular models were applied to characterize this interaction by means of co-immunoprecipitation of tagged proteins and immuno-fluorescence staining. The occurrence of the interaction at Smad2 driven transcriptomic complexes was studied by means of DNA-pull-down and chromatin immunoprecipitation (ChIP), while its effects were assessed by protein over-expression and siRNA applied into a TGFβ-dependent reporter gene assay.ResultsThe interaction was confirmed and observed to be enhanced upon Smad2 acetylation, a known feature of active and nuclear Smad2. However, Sirt1 did not play a major role in Smad2 deacetylation. Anti-Sirt1 ChIP showed increased recovery of promoter regions corresponding to Smad2-driven genes after TGFβ-stimulation, while its occurrence at Smad2-dependent transcriptomic complexes on DNA was found to effectively modulate gene expression.ConclusionsSirt1 presence on Smad2-driven TGFβ-dependent regulatory elements was detected and found to increase after TGFβ treatment. Moreover, Sirt1 overexpression resulted in a decrease of the activity of a Smad2-driven TGFβ-dependent reporter gene, while Sirt1 interference increased its activity. This would confirm the relevance of the discovered Sirt1-Smad2 interaction for the regulation of TGFβ-dependent gene transcription.Electronic supplementary materialThe online version of this article (10.1186/s12964-017-0205-y) contains supplementary material, which is available to authorized users.

Protective mechanism of SIRT1 on Hcy‐induced atrial fibrosis mediated by TRPC3

“…Although TRPC3 is well‐established to play a central role in transducing cardiac fibrosis signalling, the mechanisms underlying its signal transduction function remain poorly understood. To address this issue, researchers have identified SIRT1 as a modulator of the transcription of TGF‐β‐dependent genes, which participate in the process of fibrosis . In a preliminary experiment, we found that Hcy could increase the level of TRPC3; nevertheless, the protein level of SIRT1 was decreased (Figure A).…”

“…To address this issue, researchers have identified SIRT1 as a modulator of the transcription of TGF-β-dependent genes, which participate in the process of fibrosis. 21,22 In a preliminary experiment, we found that Hcy could increase the level of TRPC3; nevertheless, the protein level of SIRT1 was decreased ( Figure 5A). SIRT1 could be a potential TRPC3interacting partner, as both regulate the process of cardiac fibrosis in synergy.…”

Role of TGF-β in Skin Chronic Wounds: A Keratinocyte Perspective

“…Moreover, co-receptor molecules with the ability to modulate and which bind to TβRs have been described [18]. Furthermore, downstream to TβRs activation, intracellular transduction of the signal shows, again, possibilities for modulation by means of post-translational modifications [19,20] or the regulation of protein levels of the factors participating on either Smad-reliant canonical signaling [21] or MAP-kinase-dependent non-canonical signaling [22]. In that sense, the aspects of TGF-β signaling intracellular transduction and the crosstalk with other pathways have been the subject of extensive reviews [23][24][25][26].…”

The signaling mechanism of Hcy-induced atrial fibrosis mediated by TRPC3

“…Although TRPC3 is well established to play a central role in transducing cardiac fibrosis signalling [21] , the mechanisms underlying its signal transduction function still remain poorly understood, especially when TRPC3 also regulates TGF-ß. To address this issue, researchers have identified SIRT1 as a modulator of the transcription of TGF-ß-dependent genes, which participate in the process of fibrosis [22] . In a preliminary experiment, we found that Hcy could increase the level of TRPC3;…”