TIF1gamma, nouveau régulateur négatif de la voie de signalisation du TGFbeta

The cytokine TGFbeta regulates several cellular processes such as proliferation, differentiation, migration and apoptosis, from embryonic development to adulthood. TGFbeta is well described for its tumor suppressor role through antiproliferative and proapoptotic activities, in particular in epithelial cells. During tumor progression however, TGFbeta becomes a tumor promotor, favoring angiogenesis, immune suppression and inducing the epitheliomesenchymal transition. Binding of TGFbeta ligand to its receptors activate cytoplasmic messenger Smad2 and Smad3 to complex with Smad4 and shuttle into the nucleus to regulate TGFbetatarget genes expression. Recently, TIF1gamma has been described as a new negative regulator of TGFbeta signaling, through monoubiquitination of Smad4 or direct competition with Smad4 to bind activated Smad2/3. This signaling pathway has to be finely tuned to target an action dependent on a cellular context, which is why we analyze here the regulation of functional interactions between the TGFbeta canonical signaling and TIF1gamma. In this study, we show that TIF1gamma acts as a negative regulator of Smad4 functions in TGFbetasignaling during the epithelio-mesenchymal transition and during terminal differentiation of mammary epithelial cells and lactation. We are also interested in studying TIF1gamma SUMOylation as additional level of regulation of cell response to TGFbeta. Thus we characterized four functional SUMOylation sites in TIF1gamma and we found that this post-translational modification inhibits the formation of Smads transcriptional complex and is needed to temporally restrict Smad4 residence on the promoter of TGFbetatarget genes. Our results show the critical role of TIF1gamma SUMOylation in the regulation of TGFbeta- induced epithelio-mesenchymal transition. As a conclusion, our study unveils the major role of TIF1gamma in the regulation of TGFbeta transcriptional responses. Moreover, we show that TIF1gamma requires SUMOylation to exert its repressive activity on TGFbetasignalingLe TGFbeta intervient dans la régulation de nombreux processus cellulaires comme la prolifération, la différenciation, la migration, l'apoptose, du développement embryonnaire jusqu'à la vie adulte. Le TGFbeta est aujourd'hui bien décrit pour son rôle de suppresseur de tumeur de par ses activités anti-prolifératives et pro-apoptotiques, en particulier sur les cellules épithéliales. Cependant, au cours de la progression tumorale, le TGFbeta devient un promoteur de tumeur en favorisant l'angiogenèse, l'échappement de la tumeur vis-à-vis du système immunitaire et en induisant la transition épithélio-mésenchymateuse. Après fixation du ligand TGFbeta , le complexe de récepteurs active les protéines cytoplasmiques Smad2 et Smad3 qui s'associent à Smad4 pour former le complexe transcriptionnel qui se transloque alors dans le noyau pour réguler la transcription de nombreux gènes cibles. Récemment, la protéine TIF1gamma a été décrite pour intervenir dans la régulation négative de la voie du TGFbeta , en monoubiquitinant Smad4 ou en interagissant avec Smad2/3 en compétition avec Smad4. Cette voie de signalisation devant être finement contrôlée pour cibler son action en fonction du contexte cellulaire, nous analysons ici la régulation des interactions fonctionnelles entre la voie canonique du TGFbeta et la protéine TIF1gamma. Dans cette étude, nous montrons que TIF1gamma agit comme un régulateur négatif des fonctions de Smad4 dans la voie de signalisation du TGFbeta au cours du processus de transition épithélio-mésenchymateuse et au cours de la différenciation terminale des cellules épithéliales mammaires et de la lactation. Nous étudions également la SUMOylation de TIF1gamma comme nouveau niveau de régulation de la réponse cellulaire au TGFbeta . Nous avons ainsi caractérisé les sites fonctionnels de SUMOylation de TIF1gamma et montré que cette modification post-traductionnelle inhibe la formation du complexe transcriptionnel Smad et est nécessaire pour réguler temporellement la résidence de Smad4 au niveau du promoteur de gènes cibles du TGFbeta . Nos résultats montrent le rôle important de la SUMOylation de TIF1gamma dans la régulation de la transition épithélio-mésenchymateuse induite par le TGFbeta . En conclusion, notre travail met en avant le rôle majeur de TIF1gamma dans la régulation de la réponse transcriptionnelle au TGFbeta . De plus, nous montrons que la SUMOylation de TIF1gamma est nécessaire à son activité répressive sur Smad

Dynamic Regulation of Tgf-B Signaling by Tif1γ: A Computational Approach

TIF1γ (Transcriptional Intermediary Factor 1 γ) has been implicated in Smad-dependent signaling by Transforming Growth Factor beta (TGF-β). Paradoxically, TIF1γ functions both as a transcriptional repressor or as an alternative transcription factor that promotes TGF-β signaling. Using ordinary differential-equation models, we have investigated the effect of TIF1γ on the dynamics of TGF-β signaling. An integrative model that includes the formation of transient TIF1γ-Smad2-Smad4 ternary complexes is the only one that can account for TGF-β signaling compatible with the different observations reported for TIF1γ. In addition, our model predicts that varying TIF1γ/Smad4 ratios play a critical role in the modulation of the transcriptional signal induced by TGF-β, especially for short stimulation times that mediate higher threshold responses. Chromatin immunoprecipitation analyses and quantification of the expression of TGF-β target genes as a function TIF1γ/Smad4 ratios fully validate this hypothesis. Our integrative model, which successfully unifies the seemingly opposite roles of TIF1γ, also reveals how changing TIF1γ/Smad4 ratios affect the cellular response to stimulation by TGF-β, accounting for a highly graded determination of cell fate

Matrix stiffness drives epithelial–mesenchymal transition and tumour metastasis through a TWIST1–G3BP2 mechanotransduction pathway

Matrix stiffness potently regulates cellular behavior in various biological contexts. In breast tumours, the presence of dense clusters of collagen fibrils indicates increased matrix stiffness and correlates with poor survival. It is unclear how mechanical inputs are transduced into transcriptional outputs to drive tumour progression. Here we report that TWIST1 is an essential mechano-mediator that promotes epithelial-mesenchymal transition (EMT) in response to increasing matrix stiffness. High matrix stiffness promotes nuclear translocation of TWIST1 by releasing TWIST1 from its cytoplasmic binding partner G3BP2. Loss of G3BP2 leads to constitutive TWIST1 nuclear localization and synergizes with increasing matrix stiffness to induce EMT and promote tumour invasion and metastasis. In human breast tumours, collagen fiber alignment, a marker of increasing matrix stiffness, and reduced expression of G3BP2 together predict poor survival. Our findings reveal a TWIST1-G3BP2 mechanotransduction pathway that responds to biomechanical signals from the tumour microenvironment to drive EMT, invasion, and metastasis

TIF1gamma, a new negative regulator of TGFbeta signaling

Le TGFbeta intervient dans la régulation de nombreux processus cellulaires comme la prolifération, la différenciation, la migration, l'apoptose, du développement embryonnaire jusqu'à la vie adulte. Le TGFbeta est aujourd'hui bien décrit pour son rôle de suppresseur de tumeur de par ses activités anti-prolifératives et pro-apoptotiques, en particulier sur les cellules épithéliales. Cependant, au cours de la progression tumorale, le TGFbeta devient un promoteur de tumeur en favorisant l'angiogenèse, l'échappement de la tumeur vis-à-vis du système immunitaire et en induisant la transition épithélio-mésenchymateuse. Après fixation du ligand TGFbeta , le complexe de récepteurs active les protéines cytoplasmiques Smad2 et Smad3 qui s'associent à Smad4 pour former le complexe transcriptionnel qui se transloque alors dans le noyau pour réguler la transcription de nombreux gènes cibles. Récemment, la protéine TIF1gamma a été décrite pour intervenir dans la régulation négative de la voie du TGFbeta , en monoubiquitinant Smad4 ou en interagissant avec Smad2/3 en compétition avec Smad4. Cette voie de signalisation devant être finement contrôlée pour cibler son action en fonction du contexte cellulaire, nous analysons ici la régulation des interactions fonctionnelles entre la voie canonique du TGFbeta et la protéine TIF1gamma. Dans cette étude, nous montrons que TIF1gamma agit comme un régulateur négatif des fonctions de Smad4 dans la voie de signalisation du TGFbeta au cours du processus de transition épithélio-mésenchymateuse et au cours de la différenciation terminale des cellules épithéliales mammaires et de la lactation. Nous étudions également la SUMOylation de TIF1gamma comme nouveau niveau de régulation de la réponse cellulaire au TGFbeta . Nous avons ainsi caractérisé les sites fonctionnels de SUMOylation de TIF1gamma et montré que cette modification post-traductionnelle inhibe la formation du complexe transcriptionnel Smad et est nécessaire pour réguler temporellement la résidence de Smad4 au niveau du promoteur de gènes cibles du TGFbeta . Nos résultats montrent le rôle important de la SUMOylation de TIF1gamma dans la régulation de la transition épithélio-mésenchymateuse induite par le TGFbeta . En conclusion, notre travail met en avant le rôle majeur de TIF1gamma dans la régulation de la réponse transcriptionnelle au TGFbeta . De plus, nous montrons que la SUMOylation de TIF1gamma est nécessaire à son activité répressive sur Smad4The cytokine TGFbeta regulates several cellular processes such as proliferation, differentiation, migration and apoptosis, from embryonic development to adulthood. TGFbeta is well described for its tumor suppressor role through antiproliferative and proapoptotic activities, in particular in epithelial cells. During tumor progression however, TGFbeta becomes a tumor promotor, favoring angiogenesis, immune suppression and inducing the epitheliomesenchymal transition. Binding of TGFbeta ligand to its receptors activate cytoplasmic messenger Smad2 and Smad3 to complex with Smad4 and shuttle into the nucleus to regulate TGFbetatarget genes expression. Recently, TIF1gamma has been described as a new negative regulator of TGFbeta signaling, through monoubiquitination of Smad4 or direct competition with Smad4 to bind activated Smad2/3. This signaling pathway has to be finely tuned to target an action dependent on a cellular context, which is why we analyze here the regulation of functional interactions between the TGFbeta canonical signaling and TIF1gamma. In this study, we show that TIF1gamma acts as a negative regulator of Smad4 functions in TGFbetasignaling during the epithelio-mesenchymal transition and during terminal differentiation of mammary epithelial cells and lactation. We are also interested in studying TIF1gamma SUMOylation as additional level of regulation of cell response to TGFbeta. Thus we characterized four functional SUMOylation sites in TIF1gamma and we found that this post-translational modification inhibits the formation of Smads transcriptional complex and is needed to temporally restrict Smad4 residence on the promoter of TGFbetatarget genes. Our results show the critical role of TIF1gamma SUMOylation in the regulation of TGFbeta- induced epithelio-mesenchymal transition. As a conclusion, our study unveils the major role of TIF1gamma in the regulation of TGFbeta transcriptional responses. Moreover, we show that TIF1gamma requires SUMOylation to exert its repressive activity on TGFbetasignalin

Disordered protein interactions for an ordered cellular transition: Cdc2-like kinase 1 is transported to the nucleus via its Ser–Arg protein substrate

Serine-arginine (SR) proteins are essential splicing factors that promote numerous steps associated with mRNA processing and whose biological function is tightly regulated through multi-site phosphorylation. In the nucleus, the cdc2-like kinases (CLKs) phosphorylate SR proteins on their intrinsically disordered Arg-Ser (RS) domains, mobilizing them from storage speckles to the splicing machinery. The CLKs have disordered N termini that bind tightly to RS domains, enhancing SR protein phosphorylation. The N termini also promote nuclear localization of CLKs, but their transport mechanism is presently unknown. To explore cytoplasmic-nuclear transitions, several classical nuclear localization sequences in the N terminus of the CLK1 isoform were identified, but their mutation had no effect on subcellular localization. Rather, we found that CLK1 amplifies its presence in the nucleus by forming a stable complex with the SR protein substrate and appropriating its NLS for transport. These findings indicate that, along with their well-established roles in mRNA splicing, SR proteins use disordered protein-protein interactions to carry their kinase regulator from the cytoplasm to the nucleus

Mobilization of a splicing factor through a nuclear kinase-kinase complex.

The splicing of mRNA is dependent on serine-arginine (SR) proteins that are mobilized from membrane-free, nuclear speckles to the nucleoplasm by the Cdc2-like kinases (CLKs). This movement is critical for SR protein-dependent assembly of the macromolecular spliceosome. Although CLK1 facilitates such trafficking through the phosphorylation of serine-proline dipeptides in the prototype SR protein SRSF1, an unrelated enzyme known as SR protein kinase 1 (SRPK1) performs the same function but does not efficiently modify these dipeptides in SRSF1. We now show that the ability of SRPK1 to mobilize SRSF1 from speckles to the nucleoplasm is dependent on active CLK1. Diffusion from speckles is promoted by the formation of an SRPK1-CLK1 complex that facilitates dissociation of SRSF1 from CLK1 and enhances the phosphorylation of several serine-proline dipeptides in this SR protein. Down-regulation of either kinase blocks EGF-stimulated mobilization of nuclear SRSF1. These findings establish a signaling pathway that connects SRPKs to SR protein activation through the associated CLK family of kinases