Single cysteine to tyrosine transition inactivates the growth inhibitory function of Piedmontese myostatin

2002 ◽  
Vol 283 (1) ◽  
pp. C135-C141 ◽  
Author(s):  
Carole Berry ◽  
Mark Thomas ◽  
Brett Langley ◽  
Mridula Sharma ◽  
Ravi Kambadur
Keyword(s):  
Skeletal Muscle ◽  
Growth Factor ◽  
Transforming Growth Factor ◽  
Transforming Growth Factor Β ◽  
Competitive Inhibitor ◽  
Wild Type ◽  
Inhibitor Molecule ◽  
Inhibitory Function ◽  
Growth Inhibitory ◽  
Myoblast Proliferation

Myostatin, a member of the transforming growth factor-β superfamily, is a secreted growth factor that is proteolytically processed to give COOH-terminal mature myostatin and NH2-terminal latency-associated peptide in myoblasts. Piedmontese cattle are a heavy-muscled breed that express a mutated form of myostatin in which cysteine (313) is substituted with tyrosine. Here we have characterized the biology of this mutated Piedmontese myostatin. Northern and Western analyses indicate that there is increased expression of myostatin mRNA and precursor myostatin protein in the skeletal muscle of Piedmontese cattle. In contrast, a decrease in mature myostatin was observed in Piedmontese skeletal muscle. However, there is no detectable change in the circulatory levels of mature myostatin in Piedmontese cattle. Myoblast proliferation assay performed with normal and Piedmontese myostatin indicated that mature wild-type myostatin protein inhibited the proliferation of C2C12 myoblasts. Piedmontese myostatin, by contrast, failed to inhibit myoblast proliferation. In addition, when added in molar excess, Piedmontese myostatin acted as a potent “competitive inhibitor” molecule. These results indicate that, in Piedmontese myostatin, substitution of cysteine with tyrosine results in the distortion of the “cystine knot” structure and a loss of biological activity of the myostatin. This mutation also appears to affect either processing or stability of mature myostatin without altering the secretion of myostatin.

The t(3;21) Fusion Product, AML1/Evi-1, Interacts With Smad3 and Blocks Transforming Growth Factor-β–Mediated Growth Inhibition of Myeloid Cells

Blood ◽  
1998 ◽  
Vol 92 (11) ◽  
pp. 4003-4012 ◽  
Author(s):  
Mineo Kurokawa ◽  
Kinuko Mitani ◽  
Yoichi Imai ◽  
Seishi Ogawa ◽  
Yoshio Yazaki ◽  
...  
Keyword(s):  
Growth Factor ◽  
Zinc Finger ◽  
Transforming Growth Factor ◽  
Hematopoietic Cells ◽  
Myeloid Cells ◽  
Transforming Growth Factor Β ◽  
Myelogenous Leukemia ◽  
Zinc Finger Domain ◽  
Growth Inhibitory ◽  
Inhibitory Signaling

Abstract The t(3;21)(q26;q22) chromosomal translocation associated with blastic crisis of chronic myelogenous leukemia results in the formation of the AML1/Evi-1 chimeric protein, which is thought to play a causative role in leukemic transformation of hematopoietic cells. Here we show that AML1/Evi-1 represses growth-inhibitory signaling by transforming growth factor-β (TGF-β) in 32Dcl3 myeloid cells. The activity of AML1/Evi-1 to repress TGF-β signaling depends on the two separate regions of the Evi-1 portion, one of which is the first zinc finger domain. AML1/Evi-1 interacts with Smad3, an intracellular mediator of TGF-β signaling, through the first zinc finger domain, and represses the Smad3 activity, as Evi-1 does. We also show that suppression of endogenous Evi-1 in leukemic cells carrying inv(3) restores TGF-β responsiveness. Taken together, AML1/Evi-1 acts as an inhibitor of TGF-β signaling by interfering with Smad3 through the Evi-1 portion, and both AML1/Evi-1 and Evi-1 repress TGF-β–mediated growth suppression in hematopoietic cells. Thus, AML1/Evi-1 may contribute to leukemogenesis by specifically blocking growth-inhibitory signaling of TGF-β in the t(3;21) leukemia.

Transforming growth factor-β and myostatin signaling in skeletal muscle

2008 ◽  
Vol 104 (3) ◽  
pp. 579-587 ◽  
Author(s):  
Helen D. Kollias ◽  
John C. McDermott
Keyword(s):  
Skeletal Muscle ◽  
Growth Factor ◽  
Signaling Pathway ◽  
Muscle Development ◽  
Cellular Proliferation ◽  
Transforming Growth Factor ◽  
Transforming Growth Factor Β ◽  
Related Family ◽  
Cytokine Milieu

The superfamily of transforming growth factor-β (TGF-β) cytokines has been shown to have profound effects on cellular proliferation, differentiation, and growth. Recently, there have been major advances in our understanding of the signaling pathway(s) conveying TGF-β signals to the nucleus to ultimately control gene expression. One tissue that is potently influenced by TGF-β superfamily signaling is skeletal muscle. Skeletal muscle ontogeny and postnatal physiology have proven to be exquisitely sensitive to the TGF-β superfamily cytokine milieu in various animal systems from mice to humans. Recently, major strides have been made in understanding the role of TGF-β and its closely related family member, myostatin, in these processes. In this overview, we will review recent advances in our understanding of the TGF-β and myostatin signaling pathways and, in particular, focus on the implications of this signaling pathway for skeletal muscle development, physiology, and pathology.

Phosphorylation of threonine 276 in Smad4 is involved in transforming growth factor-β-induced nuclear accumulation

2003 ◽  
Vol 285 (4) ◽  
pp. C823-C830 ◽  
Author(s):  
Bernard A. J. Roelen ◽  
Ori S. Cohen ◽  
Malay K. Raychowdhury ◽  
Deborah N. Chadee ◽  
Ying Zhang ◽  
...  
Keyword(s):  
Growth Factor ◽  
Transforming Growth Factor ◽  
Phosphorylation Site ◽  
Transforming Growth Factor Β ◽  
Nuclear Accumulation ◽  
Mutant Protein ◽  
Wild Type ◽  
Type Protein ◽  
Wild Type Protein

Smad4, the common Smad, is central for transforming growth factor (TGF)-β superfamily ligand signaling. Smad4 has been shown to be constitutively phosphorylated (Nakao A, Imamura T, Souchelnytskyi S, Kawabata M, Ishisaki A, Oeda E, Tamaki K, Hanai J, Heldin C-H, Miyazono K, and ten Dijke P. EMBO J 16: 5353-5362, 1997), but the site(s) of phosphorylation, the kinase(s) that performs this phosphorylation, and the significance of the phosphorylation of Smad4 are currently unknown. This report describes the identification of a consensus ERK phosphorylation site in the linker region of Smad4 at Thr276. Our data show that ERK can phosphorylate Smad4 in vitro but not Smad4 with mutated Thr276. Flag-tagged Smad4-T276A mutant protein accumulates less efficiently in the nucleus after stimulation by TGF-β and is less efficient in generating a transcriptional response than Smad4 wild-type protein. Tryptic phosphopeptide mapping identified a phosphopeptide in Smad4 wild-type protein that was absent in phosphorylated Smad4-T276A mutant protein. Our results suggest that MAP kinase can phosphorylate Thr276 of Smad4 and that phosphorylation can lead to enhanced TGF-β-induced nuclear accumulation and, as a consequence, enhanced transcriptional activity of Smad4.

scholarly journals Local versus systemic control of bone and skeletal muscle mass by components of the transforming growth factor-β signaling pathway

2021 ◽  
Vol 118 (33) ◽  
pp. e2111401118
Author(s):  
Yewei Liu ◽  
Adam Lehar ◽  
Renata Rydzik ◽  
Harshpreet Chandok ◽  
Yun-Sil Lee ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Growth Factor ◽  
Muscle Mass ◽  
Transforming Growth Factor ◽  
Critical Role ◽  
Transforming Growth Factor Β ◽  
The Body ◽  
Dependent Manner ◽  
Bone Homeostasis ◽  
Mineral Density

Skeletal muscle and bone homeostasis are regulated by members of the myostatin/GDF-11/activin branch of the transforming growth factor-β superfamily, which share many regulatory components, including inhibitory extracellular binding proteins and receptors that mediate signaling. Here, we present the results of genetic studies demonstrating a critical role for the binding protein follistatin (FST) in regulating both skeletal muscle and bone. Using an allelic series corresponding to varying expression levels of endogenous Fst, we show that FST acts in an exquisitely dose-dependent manner to regulate both muscle mass and bone density. Moreover, by employing a genetic strategy to target Fst expression only in the posterior (caudal) region of the animal, we show that the effects of Fst loss are mostly restricted to the posterior region, implying that locally produced FST plays a much more important role than circulating FST with respect to regulation of muscle and bone. Finally, we show that targeting receptors for these ligands specifically in osteoblasts leads to dramatic increases in bone mass, with trabecular bone volume fraction being increased by 12- to 13-fold and bone mineral density being increased by 8- to 9-fold in humeri, femurs, and lumbar vertebrae. These findings demonstrate that bone, like muscle, has an enormous inherent capacity for growth that is normally kept in check by this signaling system and suggest that the extent to which this regulatory mechanism may be used throughout the body to regulate tissue mass may be more significant than previously appreciated.

Stimulation of DNA Synthesis by Interferon and Transforming Growth Factor β in EL2 Rat Fibroblasts. Role of This Effect on the Expression of an EL2-Transformed Phenoptype

1988 ◽  
Vol 1 (3) ◽  
pp. 185-191
Author(s):  
P. Di Francesco ◽  
E. Liboi ◽  
G. Febbraro ◽  
C. Favalli
Keyword(s):  
Growth Factor ◽  
Dna Synthesis ◽  
Transforming Growth Factor ◽  
Transforming Growth Factor Β ◽  
Cellular Events ◽  
Growth Inhibitory ◽  
Fibroblast Line ◽  
Epidermal Growth ◽  
Rat Fibroblasts ◽  
Malignant Conversion

The proliferation of normal cells is regulated by both positive and negative growth-effects, induced by growth-stimulatory factors [e.g., Platelet Derived Growth Factor (PDGF), Fibroblast Growth Factor (FGF), Epidermal Growth Factor (EGF)] and growth-inhibitory factors [e.g., Interferons (IFNs), Transforming Growth Factor β (TGFβ), Tumor Necrosis Factor (TNF)]. To investigate these effects, we analyzed DNA synthesis in an euploid fibroblast line (EL2), recently isolated from rat embryo, stimulated with EGF and treated with TGFβ and IFN respectively. Our results show that, in opposition to what is described for other fibroblast lines, in EL2 cells the treatment with appropriate concentrations of TGFβ or IFN did not inhibit the [3H]-thymidine incorporation induced by EGF, but, on the contrary, EGF-induced DNA synthesis was greatly stimulated by the presence of TGFβ or IFN in our cell cultures. The implications of these findings for elucidating the cellular events leading to the malignant conversion are discussed.

scholarly journals Protein tyrosine phosphatase-α amplifies transforming growth factor-β-dependent profibrotic signaling in lung fibroblasts

2020 ◽  
Vol 319 (2) ◽  
pp. L294-L311 ◽  
Author(s):  
Yael Aschner ◽  
Meghan Nelson ◽  
Matthew Brenner ◽  
Helen Roybal ◽  
Keriann Beke ◽  
...  
Keyword(s):  
Growth Factor ◽  
Epithelial Cells ◽  
Pulmonary Fibrosis ◽  
Protein Tyrosine Phosphatase ◽  
Transforming Growth Factor ◽  
Tyrosine Phosphatase ◽  
Transforming Growth Factor Β ◽  
Lung Fibroblasts ◽  
Type Ii ◽  
Wild Type

Idiopathic pulmonary fibrosis (IPF) is a progressive, often fatal, fibrosing lung disease for which treatment remains suboptimal. Fibrogenic cytokines, including transforming growth factor-β (TGF-β), are central to its pathogenesis. Protein tyrosine phosphatase-α (PTPα) has emerged as a key regulator of fibrogenic signaling in fibroblasts. We have reported that mice globally deficient in PTPα ( Ptpra−/−) were protected from experimental pulmonary fibrosis, in part via alterations in TGF-β signaling. The goal of this study was to determine the lung cell types and mechanisms by which PTPα controls fibrogenic pathways and whether these pathways are relevant to human disease. Immunohistochemical analysis of lungs from patients with IPF revealed that PTPα was highly expressed by mesenchymal cells in fibroblastic foci and by airway and alveolar epithelial cells. To determine whether PTPα promotes profibrotic signaling pathways in lung fibroblasts and/or epithelial cells, we generated mice with conditional (floxed) Ptpra alleles ( Ptpraf/f). These mice were crossed with Dermo1-Cre or with Sftpc-CreERT2 mice to delete Ptpra in mesenchymal cells and alveolar type II cells, respectively. Dermo1-Cre/ Ptpraf/f mice were protected from bleomycin-induced pulmonary fibrosis, whereas Sftpc-CreERT2 /Ptpraf/f mice developed pulmonary fibrosis equivalent to controls. Both canonical and noncanonical TGF-β signaling and downstream TGF-β-induced fibrogenic responses were attenuated in isolated Ptpra−/− compared with wild-type fibroblasts. Furthermore, TGF-β-induced tyrosine phosphorylation of TGF-β type II receptor and of PTPα were attenuated in Ptpra−/− compared with wild-type fibroblasts. The phenotype of cells genetically deficient in PTPα was recapitulated with the use of a Src inhibitor. These findings suggest that PTPα amplifies profibrotic TGF-β-dependent pathway signaling in lung fibroblasts.

Transforming Growth Factor-β Isoform Expression in the Perisutural Tissues of Craniosynostotic Rabbits

2004 ◽  
Vol 41 (4) ◽  
pp. 392-402 ◽  
Author(s):  
Elyane Poisson ◽  
James J. Sciote ◽  
Richard Koepsel ◽  
Gregory M. Cooper ◽  
Lynne A. Opperman ◽  
...  
Keyword(s):  
Growth Factor ◽  
Transforming Growth Factor ◽  
Expression Patterns ◽  
Rabbit Model ◽  
Transforming Growth Factor Β ◽  
Differential Staining ◽  
Wild Type ◽  
Opposite Pattern ◽  
Suture Fusion ◽  
Tgf Β1

Objective To describe the expression patterns of the various transforming growth factor-β (Tgf-β) isoforms, known to be involved in suture development, in the perisutural tissues of rabbits with naturally occurring craniosynostosis and relate such differential expression to the pathogenesis of premature suture fusion. Method Twenty-one coronal sutures were harvested from six wild-type control New Zealand White rabbits and five rabbits with familial coronal suture synostosis at 25 days of age for histomorphometric and immunohistochemical analyses. Tgf-β isoform immunoreactivity was assessed using indirect immunoperoxidase procedures with specific antibodies. Results Synostosed sutures had significantly (p < .01) greater bone area and relatively more osteoblasts and osteocytes in the osteogenic fronts, compared with wild-type sutures. Tgf-β isoform immunoreactivity showed differential staining patterns between wild-type and synostosed perisutural tissues. In wild-type sutures, Tgf-β1 and Tgf-β3 immunoreactivity was significantly (p < .001) greater than Tgf-β2 staining in all perisutural tissues. In synostosed sutures, the opposite pattern was observed, with Tgf-β2 immunoreactivity significantly (p < .001) greater than Tgf-β1 and Tgf-β3 in the osteogenic fronts, dura mater, and periosteum. Conclusions Findings from this study suggest that an overexpression of Tgf-β2, either in isolation or in association with an underexpression of Tgf-β1 and Tgf-β3, may be related to premature suture fusion (craniosynostosis) in this pathological rabbit model. These abnormal expression patterns may be involved in premature suture fusion either through increased cell proliferation, decreased apoptosis of the osteoblasts or both at the osteogenic fronts.

scholarly journals αB-crystallin regulation of angiogenesis by modulation of VEGF

Blood ◽  
2010 ◽  
Vol 115 (16) ◽  
pp. 3398-3406 ◽  
Author(s):  
Satoru Kase ◽  
Shikun He ◽  
Shozo Sonoda ◽  
Mizuki Kitamura ◽  
Christine Spee ◽  
...  
Keyword(s):  
Growth Factor ◽  
Protein Expression ◽  
Choroidal Neovascularization ◽  
Transforming Growth Factor ◽  
Retinal Pigment ◽  
Small Heat Shock Protein ◽  
Transforming Growth Factor Β ◽  
Wild Type ◽  
Rpe Cells ◽  
Αb Crystallin

Abstract αB-crystallin is a chaperone belonging to the small heat shock protein family. Herein we show attenuation of intraocular angiogenesis in αB-crystallin knockout (αB-crystallin−/−) mice in 2 models of intraocular disease: oxygen-induced retinopathy and laser-induced choroidal neovascularization. Vascular endothelial growth factor A (VEGF-A) mRNA and hypoxia inducible factor-1α protein expression were induced during retinal angiogenesis, but VEGF-A protein expression remained low in αB-crystallin−/− retina versus wild-type mice, whereas VEGF-R2 expression was not affected. Both αB-crystallin and its phosphorylated serine59 formwere expressed, and immunoprecipitation revealed αB-crystallin binding to VEGF-A but not transforming growth factor-β in cultured retinal pigment epithelial (RPE) cells. αB-crystallin and VEGF-A are colocalized in the endoplasmic reticulum in RPE cells under chemical hypoxia. αB-crystallin−/− RPE showed low VEGF-A secretion under serum-starved conditions compared with wild-type cells. VEGF-A is polyubiquitinated in control and αB-crystallin siRNA treated RPE; however, mono-tetra ubiquitinated VEGF-A increases with αB-crystallin knockdown. Endothelial cell apoptosis in newly formed vessels was greater in αB-crystallin−/− than wild-type mice. Proteasomal inhibition in αB-crystallin−/− mice partially restores VEGF-A secretion and angiogenic phenotype in choroidal neovascularization. Our studies indicate an important role for αB-crystallin as a chaperone for VEGF-A in angiogenesis and its potential as a therapeutic target.

scholarly journals Transforming Growth Factor β-Induced Smad1/5 Phosphorylation in Epithelial Cells Is Mediated by Novel Receptor Complexes and Is Essential for Anchorage-Independent Growth

2008 ◽  
Vol 28 (22) ◽  
pp. 6889-6902 ◽  
Author(s):  
Amanda C. Daly ◽  
Rebecca A. Randall ◽  
Caroline S. Hill
Keyword(s):  
Growth Factor ◽  
Epithelial Cells ◽  
Transcriptional Activation ◽  
Transforming Growth Factor ◽  
Transforming Growth Factor Β ◽  
Anchorage Independent Growth ◽  
Receptor Complexes ◽  
Morphogenetic Protein ◽  
Growth Inhibitory ◽  
Simultaneous Activation

ABSTRACT Transforming growth factor β (TGF-β) signals predominantly through a receptor complex comprising ALK5 and TβRII to activate receptor-regulated Smads (R-Smads) Smad2 and Smad3. In endothelial cells, however, TGF-β can additionally activate Smad1 and Smad5. Here, we report that TGF-β also strongly induces phosphorylation of Smad1/5 in many different normal epithelial cells, epithelium-derived tumor cells, and fibroblasts. We demonstrate that TβRII and ALK5, as well as ALK2 and/or ALK3, are required for TGF-β-induced Smad1/5 phosphorylation. We show that the simultaneous activation of the R-Smads Smad2/3 and Smad1/5 by TGF-β results in the formation of mixed R-Smad complexes, containing, for example, phosphorylated Smad1 and Smad2. The prevalence of these mixed R-Smad complexes explains why TGF-β-induced Smad1/5 phosphorylation does not result in transcriptional activation via bone morphogenetic protein (BMP)-responsive elements, which bind activated Smad1/5-Smad4 complexes that are induced by BMP stimulation. Thus, TGF-β induces two parallel pathways: one signaling via Smad2-Smad4 or Smad3-Smad4 complexes and the other signaling via mixed R-Smad complexes. Finally, we assess the function of the novel arm of TGF-β signaling and show that TGF-β-induced Smad1/5 activation is not required for the growth-inhibitory effects of TGF-β but is specifically required for TGF-β-induced anchorage-independent growth.

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