Smad7: licensed to kill beta-catenin

Elevated levels of inhibitory Smad7 are detected in several pathologic skin conditions; however the functional consequences of this expression have been unclear. A recent study shows that Smad7 overexpression in transgenic mouse epidermis at levels comparable to those seen in pathologic states is insufficient to block TGFbeta or BMP signaling, but instead produces striking phenotypes due to degradation of beta-catenin through a novel mechanism involving Smad7 and Smurf2.     

Roles of TGFbeta signaling in epidermal/appendage development

The transforming growth factor beta (TGFbeta) superfamily encompasses a number of structurally related proteins that can be divided into several subfamilies including TGFbetas, activins/inhibins and bone morphogenetic proteins (BMPs). The Smads are major intracellular mediators in transducing the signals of TGFbeta superfamily members, and are abundantly expressed in the developing epidermis and epidermal appendages. Moreover, the phenotypes of transgenic/knockout mice with altered components of the TGFbeta superfamily signaling pathway suggest that TGFbeta superfamily signaling is required for epidermal/appendage development. TGFbeta superfamily members are involved in most events during epidermal/appendage development through the TGFbeta signal transduction pathway and through cross talk with other signaling pathways. Future studies will be instrumental in defining the precise roles for TGFbeta superfamily signaling in epidermal/appendage development.     

Smad7 and Smad6 differentially modulate transforming growth factor beta -induced inhibition of embryonic lung morphogenesis

Transforming growth factors beta (TGF-beta) are known negative regulators of lung development, and excessive TGF-beta production has been noted in pulmonary hypoplasia associated with lung fibrosis. Inhibitory Smad7 was recently identified to antagonize TGF-beta family signaling by interfering with the activation of TGF-beta signal-transducing Smad complexes. To investigate whether Smad7 can regulate TGF-beta-induced inhibition of lung morphogenesis, ectopic overexpression of Smad7 was introduced into embryonic mouse lungs in culture using a recombinant adenovirus containing Smad7 cDNA. Although exogenous TGF-beta efficiently reduced epithelial lung branching morphogenesis in control virus-infected lung culture, TGF-beta-induced branching inhibition was abolished after epithelial transfer of the Smad7 gene into lungs in culture. Smad7 also prevented TGF-beta-mediated down-regulation of surfactant protein C gene expression, a marker of bronchial epithelial differentiation, in cultured embryonic lungs. Moreover, we found that Smad7 transgene expression blocked Smad2 phosphorylation induced by exogenous TGF-beta ligand in lung culture, indicating that Smad7 exerts its inhibitory effect on both lung growth and epithelial cell differentiation through modulation of TGF-beta pathway-restricted Smad activity. However, the above anti-TGF-beta signal transduction effects were not observed in cultured embryonic lungs with Smad6 adenoviral gene transfer, suggesting that Smad7 and Smad6 differentially regulate TGF-beta signaling in developing lungs. Our data therefore provide direct evidence that Smad7, but not Smad6, prevents TGF-beta-mediated inhibition of both lung branching morphogenesis and cytodifferentiation, establishing the mechanistic basis for Smad7 as a novel target to ameliorate aberrant TGF-beta signaling during lung development, injury, and repair.     

Antagonistic effects of Smad2 versus Smad7 are sensitive to their expression level during tooth development

Members of the transforming growth factor-beta (TGF-beta) superfamily regulate cell proliferation, differentiation, and apoptosis, controlling the development and maintenance of most tissues. TGF-beta signal is transmitted through the phosphorylation of Smad proteins by TGF-beta receptor serine/threonine kinase. During early tooth development, TGF-beta inhibits proliferation of enamel organ epithelial cells but the underlying molecular mechanisms are largely unknown. Here we tested the hypothesis that antagonistic effects between Smad2 and Smad7 regulate TGF-beta signaling during tooth development. Attenuation of Smad2 gene expression resulted in significant advancement of embryonic tooth development with increased proliferation of enamel organ epithelial cells, while attenuation of Smad7 resulted in significant inhibition of embryonic tooth development with increased apoptotic activity within enamel organ epithelium. These findings suggest that different Smads may have differential activities in regulating TGF-beta-mediated cell proliferation and death. Furthermore, functional haploinsufficiency of Smad2, but not Smad3, altered TGF-beta-mediated tooth development. The results indicate that Smads are critical factors in orchestrating TGF-beta-mediated gene regulation during embryonic tooth development. The effectiveness of TGF-beta signaling is highly sensitive to the level of Smad gene expression.     

Smad7 is required for the development and function of the heart

Transforming growth factor-beta (TGF-beta) family members, including TGF-betas, activins, and bone morphogenetic proteins, exert diverse biological activities in cell proliferation, differentiation, apoptosis, embryonic development, and many other processes. These effects are largely mediated by Smad proteins. Smad7 is a negative regulator for the signaling of TGF-beta family members. Dysregulation of Smad7 is associated with pathogenesis of a variety of human diseases. However, the in vivo physiological roles of Smad7 have not been elucidated due to the lack of a mouse model with significant loss of Smad7 function. Here we report generation and initial characterization of Smad7 mutant mice with targeted deletion of the indispensable MH2 domain. The majority of Smad7 mutant mice died in utero due to multiple defects in cardiovascular development, including ventricular septal defect and non-compaction, as well as outflow tract malformation. The surviving adult Smad7 mutant mice had impaired cardiac functions and severe arrhythmia. Further analyses suggest that Smad2/3 phosphorylation was elevated in atrioventricular cushion in the heart of Smad7 mutant mice, accompanied by increased apoptosis in this region. Taken together, these observations pinpoint an important role of Smad7 in the development and function of the mouse heart in vivo.     

Axin is a scaffold protein in TGF-beta signaling that promotes degradation of Smad7 by Arkadia

TGF-beta signaling involves a wide array of signaling molecules and multiple controlling events. Scaffold proteins create a functional proximity of signaling molecules and control the specificity of signal transduction. While many components involved in the TGF-beta pathway have been elucidated, little is known about how those components are coordinated by scaffold proteins. Here, we show that Axin activates TGF-beta signaling by forming a multimeric complex consisting of Smad7 and ubiquitin E3 ligase Arkadia. Axin depends on Arkadia to facilitate TGF-beta signaling, as their small interfering RNAs reciprocally abolished the stimulatory effect on TGF-beta signaling. Specific knockdown of Axin or Arkadia revealed that Axin and Arkadia cooperate with each other in promoting Smad7 ubiquitination. Pulse-chase experiments further illustrated that Axin significantly decreased the half-life of Smad7. Axin also induces nuclear export of Smad7. Interestingly, Axin associates with Arkadia and Smad7 independently of TGF-beta signal, in contrast to its transient association with inactive Smad3. However, coexpression of Wnt-1 reduced Smad7 ubiquitination by downregulating Axin levels, underscoring the importance of Axin as an intrinsic regulator in TGF-beta signaling.     

Threonine 41 in beta-catenin serves as a key phosphorylation relay residue in beta-catenin degradation

Beta-catenin phosphorylation at serine 45 (Ser45), threonine 41 (Thr41), Ser37, and Ser33 is critical for beta-catenin degradation, and regulation of beta-catenin phosphorylation is a central part of the canonical Wnt signaling pathway. Beta-catenin mutations at Ser45, Thr41, Ser37, and Ser33 perturb beta-catenin degradation and are frequently found in cancers. It is established that Ser45 phosphorylation by casein kinase I (CKI) initiates phosphorylation at Thr41, Ser37, and Ser33 by glycogen synthase kinase 3 (GSK3) and that phosphorylated Ser37 and Ser33 are recognized by the F-box protein beta-TrCP, a component of a ubiquitin ligase complex that mediates beta-catenin degradation. While the roles of Ser45, Ser37, and Ser33 are well documented, the function of Thr41 remains less defined. Here we show that Thr41 strictly acts as a phosphorylation relay residue and that the Ser-X-X-X-Ser (X is any amino acid) motif is obligatory for beta-catenin phosphorylation by GSK3. Beta-catenin phosphorylation/degradation and its regulation by Wnt can occur normally in the absence of Thr41 as long as the Ser-X-X-X-Ser motif/spacing is preserved. These results suggest that Thr41 functions to bridge sequential phosphorylation from Ser45 to Ser37 and provide further insights into the discrete steps and logic in beta-catenin phosphorylation-degradation.     

Compartments and appendage development in Drosophila

The appendages of Drosophila develop from the imaginal discs. During the extensive growth of these discs cell lineages are for the most part unfixed, suggesting a strong role for cell-cell interactions in controlling the final pattern of differentiation. However, during early and middle stages of development, discs are subdivided by strict lineage restrictions into a small number of spatially distinct compartments. These compartments appear to be maintained by stably inheriting states of gene expression; the compartmentspecific expression of two such ‘selector’ - like genes, engrailed and apterous, are critical for anterior-posterior and dorso-ventral compartmentalization, respectively. Recent work suggests that one purpose of compartmentalization is to establish regions of specialized cells near compartment boundaries via intercompartmental induction, using molecules like the hedgehog protein. Thus, compartments can act as organizing centers for patterning within compartments. Evidence for non-compartmental patterning mechanisms will also be discussed.     

TNF superfamily in skin appendage development

The development of skin appendages such as hairs, teeth, and mammary glands is regulated by signaling molecules of the Wnt, FGF, TGFbeta, and Hedgehog pathways. Last decade has also revealed a pivotal role for the TNF family ligand ectodysplasin (Eda) in multiple steps of epithelial appendage morphogenesis, from initiation to differentiation. Surprisingly, other members of the TNF superfamily such as Rank ligand, lymphotoxins, and TNF have recently been linked with specific aspects of skin appendage biology including branching of the mammary gland, hair shaft formation, and hair follicle cycling. This review focuses on the novel discoveries of Eda and other TNF related cytokines in skin appendage development made since the previous review on this topic in Cytokine and Growth Factor reviews in 2003.