Sirt1 interaction with active Smad2 modulates transforming growth factor-β regulated transcription
Abstract Background The 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-transla...
| Main Authors: | , , , |
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| Format: | Article |
| Language: | English |
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BMC
2017-11-01
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| Series: | Cell Communication and Signaling |
| Subjects: | |
| Online Access: | http://link.springer.com/article/10.1186/s12964-017-0205-y |
| _version_ | 1818518308541956096 |
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| author | Eva María García-Vizcaíno Sergio Liarte José Luis Alonso-Romero Francisco José Nicolás |
| author_facet | Eva María García-Vizcaíno Sergio Liarte José Luis Alonso-Romero Francisco José Nicolás |
| author_sort | Eva María García-Vizcaíno |
| collection | DOAJ |
| description | Abstract Background The 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. Methods A 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. Results The 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. Conclusions Sirt1 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. |
| first_indexed | 2024-12-11T01:08:33Z |
| format | Article |
| id | doaj.art-3e43f93adeb2462da7a50afcebadc69c |
| institution | Directory Open Access Journal |
| issn | 1478-811X |
| language | English |
| last_indexed | 2024-12-11T01:08:33Z |
| publishDate | 2017-11-01 |
| publisher | BMC |
| record_format | Article |
| series | Cell Communication and Signaling |
| spelling | doaj.art-3e43f93adeb2462da7a50afcebadc69c2022-12-22T01:26:07ZengBMCCell Communication and Signaling1478-811X2017-11-0115111710.1186/s12964-017-0205-ySirt1 interaction with active Smad2 modulates transforming growth factor-β regulated transcriptionEva María García-Vizcaíno0Sergio Liarte1José Luis Alonso-Romero2Francisco José Nicolás3Laboratorio de Oncología Molecular y TGFβ, Instituto Murciano de Investigaciones Biosanitarias ArrixacaLaboratorio de Oncología Molecular y TGFβ, Instituto Murciano de Investigaciones Biosanitarias ArrixacaServicio de Oncología, Hospital Clínico Universitario Virgen de la ArrixacaLaboratorio de Oncología Molecular y TGFβ, Instituto Murciano de Investigaciones Biosanitarias ArrixacaAbstract Background The 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. Methods A 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. Results The 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. Conclusions Sirt1 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.http://link.springer.com/article/10.1186/s12964-017-0205-ySirt1TGFβ-signalingTumor transformationProtein interactionGene transcription regulation |
| spellingShingle | Eva María García-Vizcaíno Sergio Liarte José Luis Alonso-Romero Francisco José Nicolás Sirt1 interaction with active Smad2 modulates transforming growth factor-β regulated transcription Cell Communication and Signaling Sirt1 TGFβ-signaling Tumor transformation Protein interaction Gene transcription regulation |
| title | Sirt1 interaction with active Smad2 modulates transforming growth factor-β regulated transcription |
| title_full | Sirt1 interaction with active Smad2 modulates transforming growth factor-β regulated transcription |
| title_fullStr | Sirt1 interaction with active Smad2 modulates transforming growth factor-β regulated transcription |
| title_full_unstemmed | Sirt1 interaction with active Smad2 modulates transforming growth factor-β regulated transcription |
| title_short | Sirt1 interaction with active Smad2 modulates transforming growth factor-β regulated transcription |
| title_sort | sirt1 interaction with active smad2 modulates transforming growth factor β regulated transcription |
| topic | Sirt1 TGFβ-signaling Tumor transformation Protein interaction Gene transcription regulation |
| url | http://link.springer.com/article/10.1186/s12964-017-0205-y |
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