Tif1 is essential for the terminal differentiation of mammary alveolar epithelial cells and for lactation through SMAD4 inhibition

Development (Impact Factor: 6.46). 11/2012; 140(1). DOI: 10.1242/dev.085068
Source: PubMed


Transforming growth factor β (TGFβ) is widely recognised as an important factor that regulates many steps of normal mammary gland (MG) development, including branching morphogenesis, functional differentiation and involution. Tif1γ has previously been reported to temporally and spatially control TGFβ signalling during early vertebrate development by exerting negative effects over SMAD4 availability. To evaluate the contribution of Tif1 γ to MG development, we developed a Cre/LoxP system to specifically invalidate the Tif1g gene in mammary epithelial cells in vivo. Tif1g-null mammary gland development appeared to be normal and no defects were observed during the lifespan of virgin mice. However, a lactation defect was observed in mammary glands of Tif1g-null mice. We demonstrate that Tif1 γ is essential for the terminal differentiation of alveolar epithelial cells at the end of pregnancy and to ensure lactation. Tif1 γ appears to play a crucial role in the crosstalk between TGFβ and prolactin pathways by negatively regulating both PRL receptor expression and STAT5 phosphorylation, thereby impairing the subsequent transactivation of PRL target genes. Using HC11 cells as a model, we demonstrate that the effects of Tif1g knockdown on lactation depend on both SMAD4 and TGFβ. Interestingly, we found that the Tif1γ expression pattern in mammary epithelial cells is almost symmetrically opposite to that described for TGFβ. We propose that Tif1γ contributes to the repression of TGFβ activity during late pregnancy and prevents lactation by inhibiting SMAD4.

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Available from: Germain Gillet, Apr 16, 2015
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    • "Ubiquitylationmediated degradation of Smad4 can be antagonized by the FAM/ USP9x de-ubiquitylase, thus restoring Smad4 function (Dupont et al., 2009). Results from others as well as our own have shown that TIF1c is a potent inhibitor of Smad4 functions during TGFbinduced EMT (Hesling et al., 2011), terminal differentiation of mammary alveolar epithelial cells and lactation (Hesling et al., 2013) and during specification of the ectoderm in Xenopus embryos (Dupont et al., 2005). Moreover the negative regulation of Smad4 by TIF1c is essential to achieve proper dosage of Nodal responsiveness in mouse embryos (Morsut et al., 2010). "
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    ABSTRACT: TIF1γ, a new actor of TGFβ signaling, inhibits the Smad4-mediated TGFβ response by interaction with Smad2/3 or ubiquitination of Smad4. We have shown that TIF1γ participates in TGFβ signaling as a negative regulator of Smad4 during the TGFβ-induced epithelial-to-mesenchymal transition in mammary epithelial cells and during terminal differentiation of mammary alveolar epithelial cells and lactation. We demonstrate here that TIF1γ is sumoylated and interacts with Ubc9, the only known SUMO-conjugating enzyme. Four functional sumoylation sites lie within the middle domain of TIF1γ, the Smad interaction domain. We show that a sumoylation-defective TIF1γ mutant significantly reduces TIF1γ inhibition of Smad complexes and that of the Smad-mediated TGFβ transcriptional response. Moreover, chromatin immunoprecipitation experiments indicate that TIF1γ sumoylation is required to limit Smad4 binding on the PAI-1 TGFβ target gene promoter. Ectopic expression of TIF1γ in mammary epithelial cells inhibits TGFβ-induced EMT, an effect relieved by expression of non-sumoylated TIF1γ. Taken together, our results identify a new TGFβ regulatory layer, whereby sumoylation strengthens the TIF1γ repressive action on canonical TGFβ signaling.
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    ABSTRACT: Members of the Transforming Growth Factor-β (TGF-β) family of secreted proteins are present in all multicellular animals. TGF-β proteins are versatile intercellular signaling molecules that orchestrate cell fate decisions during development and maintain homeostasis in adults. The Smad family of signal transducers implements TGF-β signals in responsive cells. Given the ability of TGF-β ligands to induce dramatic responses in target cells, numerous regulatory mechanisms exist to prevent unintended consequences. Here we review new reports of extracellular and post-translational regulation in Drosophila and vertebrates. Extracellular topics include the regulation of TGF-β signaling range and the coordination between tissue morphogenesis and TGF-β signaling. Post-translational topics include the regulation of TGF-β signal transduction by Gsk3-β phosphorylation of Smads and by cycles of Smad mono- and deubiquitylation. Extension of the ubiquitylation data to the Hippo pathway is also discussed.
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