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Tubita A, Menconi A, Lombardi Z, Tusa I, Esparís-Ogando A, Pandiella A, Gamberi T, Stecca B, Rovida E. Latent-Transforming Growth Factor β-Binding Protein 1/Transforming Growth Factor β1 Complex Drives Antitumoral Effects upon ERK5 Targeting in Melanoma. THE AMERICAN JOURNAL OF PATHOLOGY 2024; 194:1581-1591. [PMID: 38705382 DOI: 10.1016/j.ajpath.2024.03.015] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 09/15/2023] [Revised: 03/14/2024] [Accepted: 03/27/2024] [Indexed: 05/07/2024]
Abstract
Melanoma is the deadliest skin cancer, with a poor prognosis in advanced stages. While available treatments have improved survival, long-term benefits are still unsatisfactory. The mitogen-activated protein kinase extracellular signal-regulated kinase 5 (ERK5) promotes melanoma growth, and ERK5 inhibition determines cellular senescence and the senescence-associated secretory phenotype. Here, latent-transforming growth factor β-binding protein 1 (LTBP1) mRNA was found to be up-regulated in A375 and SK-Mel-5 BRAF V600E melanoma cells after ERK5 inhibition. In keeping with a key role of LTBP1 in regulating transforming growth factor β (TGF-β), TGF-β1 protein levels were increased in lysates and conditioned media of ERK5-knockdown (KD) cells, and were reduced upon LTBP1 KD. Both LTBP1 and TGF-β1 proteins were increased in melanoma xenografts in mice treated with the ERK5 inhibitor XMD8-92. Moreover, treatment with conditioned media from ERK5-KD melanoma cells reduced cell proliferation and invasiveness, and TGF-β1-neutralizing antibodies impaired these effects. In silico data sets revealed that higher expression levels of both LTBP1 and TGF-β1 mRNA were associated with better overall survival of melanoma patients. Increased LTBP1 or TGF-β1 expression played a beneficial role in patients treated with anti-PD1 immunotherapy, making a possible immunosuppressive role of LTBP1/TGF-β1 unlikely upon ERK5 inhibition. This study, therefore, identifies additional desirable effects of ERK5 targeting, providing evidence of an ERK5-dependent tumor-suppressive role of TGF-β in melanoma.
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Affiliation(s)
- Alessandro Tubita
- Department of Clinical and Experimental Biomedical Sciences, University of Florence, Florence, Italy
| | - Alessio Menconi
- Department of Clinical and Experimental Biomedical Sciences, University of Florence, Florence, Italy
| | - Zoe Lombardi
- Department of Clinical and Experimental Biomedical Sciences, University of Florence, Florence, Italy
| | - Ignazia Tusa
- Department of Clinical and Experimental Biomedical Sciences, University of Florence, Florence, Italy
| | - Azucena Esparís-Ogando
- Instituto de Biología Molecular y Celular del Cáncer (IBMCC)-Consejo Superior de Investigaciones Científicas (CSIC), Salamanca, Spain; Instituto de Investigación Biomédica de Salamanca (IBSAL), Salamanca, Spain; Centro de Investigación Biomédica en Red de Cáncer (CIBERONC), Salamanca, Spain
| | - Atanasio Pandiella
- Instituto de Biología Molecular y Celular del Cáncer (IBMCC)-Consejo Superior de Investigaciones Científicas (CSIC), Salamanca, Spain; Instituto de Investigación Biomédica de Salamanca (IBSAL), Salamanca, Spain; Centro de Investigación Biomédica en Red de Cáncer (CIBERONC), Salamanca, Spain
| | - Tania Gamberi
- Department of Clinical and Experimental Biomedical Sciences, University of Florence, Florence, Italy
| | - Barbara Stecca
- Core Research Laboratory, Institute for Cancer Research and Prevention, Florence, Italy
| | - Elisabetta Rovida
- Department of Clinical and Experimental Biomedical Sciences, University of Florence, Florence, Italy.
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Deng L, Xu G, Huang Q. Comprehensive analyses of the microRNA-messenger RNA-transcription factor regulatory network in mouse and human renal fibrosis. Front Genet 2022; 13:925097. [PMID: 36457754 PMCID: PMC9705735 DOI: 10.3389/fgene.2022.925097] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/21/2022] [Accepted: 10/14/2022] [Indexed: 09/19/2023] Open
Abstract
Objective: The aim of this study was to construct a microRNA (miRNA)-messenger RNA (mRNA)-transcription factor (TF) regulatory network and explore underlying molecular mechanisms, effective biomarkers, and drugs in renal fibrosis (RF). Methods: A total of six datasets were downloaded from Gene Expression Omnibus. "Limma" and "DESeq2" packages in R software and GEO2R were applied to identify the differentially expressed miRNAs and mRNAs (DEmiRNAs and DEmRNAs, respectively). The determination and verification of DEmiRNAs and DEmRNAs were performed through the integrated analysis of datasets from five mouse 7 days of unilateral ureteral obstruction datasets and one human chronic kidney disease dataset and the Human Protein Atlas (http://www.proteinatlas.org). Target mRNAs of DEmiRNAs and TFs were predicted by prediction databases and the iRegulon plugin in Cytoscape, respectively. A protein-protein interaction network was constructed using STRING, Cytoscape v3.9.1, and CytoNCA. Functional enrichment analysis was performed by DIANA-miRPath v3.0 and R package "clusterProfiler." A miRNA-mRNA-TF network was established using Cytoscape. Receiver operating characteristic (ROC) curve analysis was used to examine the diagnostic value of the key hub genes. Finally, the Comparative Toxicogenomics Database and Drug-Gene Interaction database were applied to identify potential drugs. Results: Here, 4 DEmiRNAs and 11 hub genes were determined and confirmed in five mouse datasets, of which Bckdha and Vegfa were further verified in one human dataset and HPA, respectively. Moreover, Bckdha and Vegfa were also predicted by miR-125a-3p and miR-199a-5p, respectively, in humans as in mice. The sequences of miR-125a-3p and miR-199a-5p in mice were identical to those in humans. A total of 6 TFs were predicted to regulate Bckdha and Vegfa across mice and humans; then, a miRNA-mRNA-TF regulatory network was built. Subsequently, ROC curve analysis showed that the area under the curve value of Vegfa was 0.825 (p = 0.002). Finally, enalapril was identified to target Vegfa for RF therapy. Conclusion: Pax2, Pax5, Sp1, Sp2, Sp3, and Sp4 together with Bckdha-dependent miR-125a-3p/Vegfa-dependent miR-199a-5p formed a co-regulatory network enabling Bckdha/Vegfa to be tightly controlled in the underlying pathogenesis of RF across mice and humans. Vegfa could act as a potential novel diagnostic marker and might be targeted by enalapril for RF therapy.
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Affiliation(s)
- Le Deng
- Department of Nephrology, The Second Affiliated Hospital of Nanchang University, Jiangxi, China
| | - Gaosi Xu
- Department of Nephrology, The Second Affiliated Hospital of Nanchang University, Jiangxi, China
| | - Qipeng Huang
- Department of Nephrology, The Fifth Affiliated Hospital of Jinan University, Heyuan, China
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Zhang J, Zhang Z, Holst S, Blöchl C, Madunic K, Wuhrer M, Ten Dijke P, Zhang T. Transforming growth factor-β challenge alters the N-, O-, and glycosphingolipid glycomes in PaTu-S pancreatic adenocarcinoma cells. J Biol Chem 2022; 298:101717. [PMID: 35151689 PMCID: PMC8914387 DOI: 10.1016/j.jbc.2022.101717] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/30/2021] [Revised: 01/27/2022] [Accepted: 01/28/2022] [Indexed: 11/13/2022] Open
Abstract
Pancreatic ductal adenocarcinoma (PDAC) is characterized by poor prognosis and high mortality. Transforming growth factor-β (TGF-β) plays a key role in PDAC tumor progression, which is often associated with aberrant glycosylation. However, how PDAC cells respond to TGF-β and the role of glycosylation therein is not well known. Here, we investigated the TGF-β-mediated response and glycosylation changes in the PaTu-8955S (PaTu-S) cell line deficient in SMA-related and MAD-related protein 4 (SMAD4), a signal transducer of the TGF-β signaling. PaTu-S cells responded to TGF-β by upregulating SMAD2 phosphorylation and target gene expression. We found that TGF-β induced expression of the mesenchymal marker N-cadherin but did not significantly affect epithelial marker E-cadherin expression. We also examined differences in N-glycans, O-glycans, and glycosphingolipid-linked glycans in PaTu-S cells upon TGF-β stimulation. TGF-β treatment primarily induced N-glycome aberrations involving elevated levels of branching, core fucosylation, and sialylation in PaTu-S cells, in agreement with TGF-β-induced changes in the expression of glycosylation-associated genes. In addition, we observed differences in O glycosylation and glycosphingolipid glycosylation profiles after TGF-β treatment, including lower levels of sialylated Tn antigen and neoexpression of globosides. Furthermore, the expression of transcription factor sex-determining region Y-related high-mobility group box 4 was upregulated upon TGF-β stimulation, and its depletion blocked TGF-β-induced N-glycomic changes. Thus, TGF-β-induced N-glycosylation changes can occur in a sex-determining region Y-related high-mobility group box 4–dependent and SMAD4-independent manner in the pancreatic PaTu-S cancer cell line. Our results open up avenues to study the relevance of glycosylation in TGF-β signaling in SMAD4-inactivated PDAC.
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Affiliation(s)
- Jing Zhang
- Oncode Institute and Department of Cell Chemical Biology, Leiden University Medical Center, Leiden, The Netherlands
| | - Zejian Zhang
- Center for Proteomics and Metabolomics, Leiden University Medical Center, Leiden, The Netherlands
| | - Stephanie Holst
- Center for Proteomics and Metabolomics, Leiden University Medical Center, Leiden, The Netherlands
| | - Constantin Blöchl
- Center for Proteomics and Metabolomics, Leiden University Medical Center, Leiden, The Netherlands; Department of Biosciences, University of Salzburg, Salzburg, Austria
| | - Katarina Madunic
- Center for Proteomics and Metabolomics, Leiden University Medical Center, Leiden, The Netherlands
| | - Manfred Wuhrer
- Center for Proteomics and Metabolomics, Leiden University Medical Center, Leiden, The Netherlands
| | - Peter Ten Dijke
- Oncode Institute and Department of Cell Chemical Biology, Leiden University Medical Center, Leiden, The Netherlands.
| | - Tao Zhang
- Center for Proteomics and Metabolomics, Leiden University Medical Center, Leiden, The Netherlands.
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Li X, Nania S, Kleiter I, Löhr JM, Heuchel RL. Targeting of Smad7 in Mesenchymal Cells Does Not Exacerbate Fibrosis During Experimental Chronic Pancreatitis. Pancreas 2021; 50:1427-1434. [PMID: 35041343 DOI: 10.1097/mpa.0000000000001951] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/10/2022]
Abstract
OBJECTIVES Transforming growth factor-β (TGF-β)-mediated accumulation of extracellular matrix proteins such as collagen I is a common feature of fibrosis. Pancreatic stellate cells play an integral role in the pathogenesis of pancreatitis, and their profibrotic ability is mainly mediated by TGF-β signaling. To specifically address the role of fibrogenic cells in experimental pancreatic fibrosis, we deleted Smad7, the main feedback inhibitor of TGF-β signaling in this cell type in mice. METHODS A mouse strain harboring a conditional knockout allele of Smad7 (Smad7fl/fl) with the tamoxifen-inducible inducible Col1a2-CreERT allele was generated and compared with wild-type mice challenged with the cerulein-based model of chronic pancreatitis. RESULTS Pancreatic stellate cells lacking Smad7 had significantly increased collagen I and fibronectin production and showed a higher activation level in vitro. Surprisingly, the fibrotic index in the pancreata of treated conditional knockout mice was only slightly increased, without statistical significance. Except for fibronectin, the expression of different extracellular matrix proteins and the numbers of fibroblasts and inflammatory cells were similar between Smad7-mutant and control mice. CONCLUSIONS There was no clear evidence that the lack of Smad7 in pancreatic stellate cells plays a major role in experimental pancreatitis, at least in the mouse model investigated here.
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Affiliation(s)
- Xuan Li
- From the Pancreas Cancer Research (PaCaRes) Lab, Department of Clinical Science, Intervention and Technology (CLINTEC), Karolinska Institutet, Stockholm, Sweden
| | - Salvatore Nania
- From the Pancreas Cancer Research (PaCaRes) Lab, Department of Clinical Science, Intervention and Technology (CLINTEC), Karolinska Institutet, Stockholm, Sweden
| | - Ingo Kleiter
- Department of Neurology, Ruhr-Universität Bochum, Bochum, Germany
| | - J-Matthias Löhr
- From the Pancreas Cancer Research (PaCaRes) Lab, Department of Clinical Science, Intervention and Technology (CLINTEC), Karolinska Institutet, Stockholm, Sweden
| | - Rainer L Heuchel
- From the Pancreas Cancer Research (PaCaRes) Lab, Department of Clinical Science, Intervention and Technology (CLINTEC), Karolinska Institutet, Stockholm, Sweden
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Dumbrava MG, Lacanlale JL, Rowan CJ, Rosenblum ND. Transforming growth factor beta signaling functions during mammalian kidney development. Pediatr Nephrol 2021; 36:1663-1672. [PMID: 32880018 DOI: 10.1007/s00467-020-04739-5] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 07/13/2020] [Revised: 07/22/2020] [Accepted: 08/04/2020] [Indexed: 12/21/2022]
Abstract
Aberrant transforming growth factor beta (TGFβ) signaling during embryogenesis is implicated in severe congenital abnormalities, including kidney malformations. However, the molecular mechanisms that underlie congenital kidney malformations related to TGFβ signaling remain poorly understood. Here, we review current understanding of the lineage-specific roles of TGFβ signaling during kidney development and how dysregulation of TGFβ signaling contributes to the pathogenesis of kidney malformation.
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Affiliation(s)
- Mihai G Dumbrava
- Program in Developmental and Stem Cell Biology, The Hospital for Sick Children, 686 Bay Street, Toronto, M5G 0A4, Canada
| | - Jon L Lacanlale
- Program in Developmental and Stem Cell Biology, The Hospital for Sick Children, 686 Bay Street, Toronto, M5G 0A4, Canada
- Department of Laboratory Medicine and Pathobiology, University of Toronto, Toronto, M5S 1A8, Canada
| | - Christopher J Rowan
- Program in Developmental and Stem Cell Biology, The Hospital for Sick Children, 686 Bay Street, Toronto, M5G 0A4, Canada
| | - Norman D Rosenblum
- Program in Developmental and Stem Cell Biology, The Hospital for Sick Children, 686 Bay Street, Toronto, M5G 0A4, Canada.
- Department of Laboratory Medicine and Pathobiology, University of Toronto, Toronto, M5S 1A8, Canada.
- Department of Physiology, University of Toronto, Toronto, M5S 1A8, Canada.
- Department of Paediatrics, University of Toronto, Toronto, M5S 1A8, Canada.
- Division of Nephrology, The Hospital for Sick Children, 555 University Avenue, Toronto, M5G 1X8, Canada.
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DiNicolantonio JJ, McCarty MF, Barroso-Aranda J, Assanga S, Lujan LML, O'Keefe JH. A nutraceutical strategy for downregulating TGFβ signalling: prospects for prevention of fibrotic disorders, including post-COVID-19 pulmonary fibrosis. Open Heart 2021; 8:openhrt-2021-001663. [PMID: 33879509 PMCID: PMC8061562 DOI: 10.1136/openhrt-2021-001663] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Accepted: 03/30/2021] [Indexed: 12/14/2022] Open
Affiliation(s)
- James J DiNicolantonio
- Preventive Cardiology, Saint Luke's Mid America Heart Institute, Kansas City, Missouri, USA
| | | | | | - Simon Assanga
- Department of Research and Postgraduate Studies in Food, University of Sonora, Sonora, Mexico
| | | | - James H O'Keefe
- University of Missouri-Kansas City, Saint Lukes Mid America Heart Institute, Kansas City, Missouri, USA
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Enhanced Intestinal TGF-β/SMAD-Dependent Signaling in Simian Immunodeficiency Virus Infected Rhesus Macaques. Cells 2021; 10:cells10040806. [PMID: 33916615 PMCID: PMC8066988 DOI: 10.3390/cells10040806] [Citation(s) in RCA: 12] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/29/2021] [Revised: 03/18/2021] [Accepted: 04/01/2021] [Indexed: 12/12/2022] Open
Abstract
Transforming growth factor-β signaling (TGF-β) maintains a balanced physiological function including cell growth, differentiation, and proliferation and regulation of immune system by modulating either SMAD2/3 and SMAD7 (SMAD-dependent) or SMAD-independent signaling pathways under normal conditions. Increased production of TGF-β promotes immunosuppression in Human Immunodeficiency Virus (HIV)/Simian Immunodeficiency Virus (SIV) infection. However, the cellular source and downstream events of increased TGF-β production that attributes to its pathological manifestations remain unknown. Here, we have shown increased production of TGF-β in a majority of intestinal CD3−CD20−CD68+ cells from acute and chronically SIV infected rhesus macaques, which negatively correlated with the frequency of jejunum CD4+ T cells. No significant changes in intestinal TGF-β receptor II expression were observed but increased production of the pSMAD2/3 protein and SMAD3 gene expression in jejunum tissues that were accompanied by a downregulation of SMAD7 protein and gene expression. Enhanced TGF-β production by intestinal CD3−CD20−CD68+ cells and increased TGF-β/SMAD-dependent signaling might be due to a disruption of a negative feedback loop mediated by SMAD7. This suggests that SIV infection impacts the SMAD-dependent signaling pathway of TGF-β and provides a potential framework for further study to understand the role of viral factor(s) in modulating TGF-β production and downregulating SMAD7 expression in SIV. Regulation of mucosal TGF-β expression by therapeutic TGF-β blockers may help to create effective antiviral mucosal immune responses.
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Di Gregorio J, Robuffo I, Spalletta S, Giambuzzi G, De Iuliis V, Toniato E, Martinotti S, Conti P, Flati V. The Epithelial-to-Mesenchymal Transition as a Possible Therapeutic Target in Fibrotic Disorders. Front Cell Dev Biol 2020; 8:607483. [PMID: 33409282 PMCID: PMC7779530 DOI: 10.3389/fcell.2020.607483] [Citation(s) in RCA: 99] [Impact Index Per Article: 19.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/17/2020] [Accepted: 11/25/2020] [Indexed: 12/11/2022] Open
Abstract
Fibrosis is a chronic and progressive disorder characterized by excessive deposition of extracellular matrix, which leads to scarring and loss of function of the affected organ or tissue. Indeed, the fibrotic process affects a variety of organs and tissues, with specific molecular background. However, two common hallmarks are shared: the crucial role of the transforming growth factor-beta (TGF-β) and the involvement of the inflammation process, that is essential for initiating the fibrotic degeneration. TGF-β in particular but also other cytokines regulate the most common molecular mechanism at the basis of fibrosis, the Epithelial-to-Mesenchymal Transition (EMT). EMT has been extensively studied, but not yet fully explored as a possible therapeutic target for fibrosis. A deeper understanding of the crosstalk between fibrosis and EMT may represent an opportunity for the development of a broadly effective anti-fibrotic therapy. Here we report the evidences of the relationship between EMT and multi-organ fibrosis, and the possible therapeutic approaches that may be developed by exploiting this relationship.
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Affiliation(s)
- Jacopo Di Gregorio
- Burnett School of Biomedical Sciences, College of Medicine, University of Central Florida, Orlando, FL, United States
| | - Iole Robuffo
- Institute of Molecular Genetics, National Research Council, Section of Chieti, Chieti, Italy
| | - Sonia Spalletta
- Department of Clinical Pathology, E. Profili Hospital, Fabriano, Ancona, Italy
| | - Giulia Giambuzzi
- Department of Medical and Oral Sciences and Biotechnologies, University “G. d’Annunzio”, Chieti, Italy
| | - Vincenzo De Iuliis
- Department of Medical and Oral Sciences and Biotechnologies, University “G. d’Annunzio”, Chieti, Italy
| | - Elena Toniato
- Department of Medical and Oral Sciences and Biotechnologies, University “G. d’Annunzio”, Chieti, Italy
| | - Stefano Martinotti
- Department of Medical and Oral Sciences and Biotechnologies, University “G. d’Annunzio”, Chieti, Italy
| | - Pio Conti
- Postgraduate Medical School, University of Chieti-Pescara, Chieti, Italy
| | - Vincenzo Flati
- Department of Biotechnological and Applied Clinical Sciences, University of L’Aquila, L’Aquila, Italy
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de Ceuninck van Capelle C, Spit M, Ten Dijke P. Current perspectives on inhibitory SMAD7 in health and disease. Crit Rev Biochem Mol Biol 2020; 55:691-715. [PMID: 33081543 DOI: 10.1080/10409238.2020.1828260] [Citation(s) in RCA: 45] [Impact Index Per Article: 9.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/12/2022]
Abstract
Transforming growth factor β (TGF-β) family members play an extensive role in cellular communication that orchestrates both early development and adult tissue homeostasis. Aberrant TGF-β family signaling is associated with a pathological outcome in numerous diseases, and in-depth understanding of molecular and cellular processes could result in therapeutic benefit for patients. Canonical TGF-β signaling is mediated by receptor-regulated SMADs (R-SMADs), a single co-mediator SMAD (Co-SMAD), and inhibitory SMADs (I-SMADs). SMAD7, one of the I-SMADs, is an essential negative regulator of the pleiotropic TGF-β and bone morphogenetic protein (BMP) signaling pathways. In a negative feedback loop, SMAD7 inhibits TGF-β signaling by providing competition for TGF-β type-1 receptor (TβRI), blocking phosphorylation and activation of SMAD2. Moreover, SMAD7 recruits E3 ubiquitin SMURF ligases to the type I receptor to promote ubiquitin-mediated proteasomal degradation. In addition to its role in TGF-β and BMP signaling, SMAD7 is regulated by and implicated in a variety of other signaling pathways and functions as a mediator of crosstalk. This review is focused on SMAD7, its function in TGF-β and BMP signaling, and its role as a downstream integrator and crosstalk mediator. This crucial signaling molecule is tightly regulated by various mechanisms. We provide an overview of the ways by which SMAD7 is regulated, including noncoding RNAs (ncRNAs) and post-translational modifications (PTMs). Finally, we discuss its role in diseases, such as cancer, fibrosis, and inflammatory bowel disease (IBD).
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Affiliation(s)
| | - Maureen Spit
- Oncode Institute and Department of Cell and Chemical Biology, Leiden University Medical Center, Leiden, the Netherlands
| | - Peter Ten Dijke
- Oncode Institute and Department of Cell and Chemical Biology, Leiden University Medical Center, Leiden, the Netherlands
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Sun Z, Cai S, Zabkiewicz C, Liu C, Ye L. Bone morphogenetic proteins mediate crosstalk between cancer cells and the tumour microenvironment at primary tumours and metastases (Review). Int J Oncol 2020; 56:1335-1351. [PMID: 32236571 DOI: 10.3892/ijo.2020.5030] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/13/2019] [Accepted: 02/28/2020] [Indexed: 11/05/2022] Open
Abstract
Bone morphogenetic proteins (BMP) are pluripotent molecules, co‑ordinating cellular functions from early embryonic and postnatal development to tissue repair, regeneration and homeostasis. They are also involved in tumourigenesis, disease progression and the metastasis of various solid tumours. Emerging evidence has indicated that BMPs are able to promote disease progression and metastasis by orchestrating communication between cancer cells and the surrounding microenvironment. The interactions occur between BMPs and epidermal growth factor receptor, hepatocyte growth factor, fibroblast growth factor, vascular endothelial growth factor and extracellular matrix components. Overall, these interactions co‑ordinate the cellular functions of tumour cells and other types of cell in the tumour to promote the growth of the primary tumour, local invasion, angiogenesis and metastasis, and the establishment and survival of cancer cells in the metastatic niche. Therefore, the present study aimed to provide an informative summary of the involvement of BMPs in the tumour microenvironment.
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Affiliation(s)
- Zhiwei Sun
- VIP‑II Division of Medical Department, Key Laboratory of Carcinogenesis and Translational Research (Ministry of Education, Beijing), Peking University Cancer Hospital and Institute, Beijing 100142, P.R. China
| | - Shuo Cai
- Cardiff China Medical Research Collaborative, Division of Cancer and Genetics, Cardiff University School of Medicine, Cardiff CF14 4XN, United Kingdom
| | - Catherine Zabkiewicz
- Cardiff China Medical Research Collaborative, Division of Cancer and Genetics, Cardiff University School of Medicine, Cardiff CF14 4XN, United Kingdom
| | - Chang Liu
- Cardiff China Medical Research Collaborative, Division of Cancer and Genetics, Cardiff University School of Medicine, Cardiff CF14 4XN, United Kingdom
| | - Lin Ye
- Cardiff China Medical Research Collaborative, Division of Cancer and Genetics, Cardiff University School of Medicine, Cardiff CF14 4XN, United Kingdom
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Andolfo I, Rosato BE, Manna F, De Rosa G, Marra R, Gambale A, Girelli D, Russo R, Iolascon A. Gain-of-function mutations in PIEZO1 directly impair hepatic iron metabolism via the inhibition of the BMP/SMADs pathway. Am J Hematol 2020; 95:188-197. [PMID: 31737919 DOI: 10.1002/ajh.25683] [Citation(s) in RCA: 40] [Impact Index Per Article: 8.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/12/2019] [Accepted: 11/13/2019] [Indexed: 02/06/2023]
Abstract
Dehydrated hereditary stomatocytosis (DHS), or xerocytosis, is an autosomal dominant hemolytic anemia. Most patients with DHS carry mutations in the PIEZO1 gene encoding a mechanosensitive cation channel. We here demonstrate that patients with DHS have low levels of hepcidin and only a slight increase of ERFE, the erythroid negative regulator of hepcidin. We demonstrated that at the physiological level, PIEZO1 activation induced Ca2+ influx and suppression of HAMP expression in primary hepatocytes. In two hepatic cellular models expressing PIEZO1 WT and two PIEZO1 gain-of-function mutants (R2456H and R2488Q), we highlight altered expression of a few genes/proteins involved in iron metabolism. Mutant cells showed increased intracellular Ca2+ compared to WT, which was correlated to increased phosphorylation of ERK1/2, inhibition of the BMP-SMADs pathway, and suppression of HAMP transcription. Moreover, the HuH7 cells, treated with PD0325901, a potent inhibitor of ERK1/2 phosphorylation, reduced the phosphorylation of ERK1/2 with the consequent increased phosphorylation of SMAD1/5/8, confirming the link between the two pathways. Another "proof of concept" for the mechanism that links PIEZO1 to HAMP regulation was obtained by mimicking PIEZO1 activation by cell Ca2+ overload, by the Ca2+ ionophore A23187. There was strong down-regulation of HAMP gene expression after this Ca2+ overload. Finally, the inhibition of PIEZO1 by GsMTx4 leads to phenotype rescue. This is the first demonstration of a direct link between PIEZO1 and iron metabolism, which defines the channel as a new hepatic iron metabolism regulator and as a possible therapeutic target of iron overload in DHS and other iron-loading anemias.
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Affiliation(s)
- Immacolata Andolfo
- Dipartimento di Medicina Molecolare e Biotecnologie MedicheUniversità degli Studi di Napoli ‘Federico II’ Naples Italy
- CEINGE, Biotecnologie Avanzate Naples Italy
| | - Barbara Eleni Rosato
- Dipartimento di Medicina Molecolare e Biotecnologie MedicheUniversità degli Studi di Napoli ‘Federico II’ Naples Italy
- CEINGE, Biotecnologie Avanzate Naples Italy
| | - Francesco Manna
- Dipartimento di Medicina Molecolare e Biotecnologie MedicheUniversità degli Studi di Napoli ‘Federico II’ Naples Italy
- CEINGE, Biotecnologie Avanzate Naples Italy
| | - Gianluca De Rosa
- Dipartimento di Medicina Molecolare e Biotecnologie MedicheUniversità degli Studi di Napoli ‘Federico II’ Naples Italy
- CEINGE, Biotecnologie Avanzate Naples Italy
| | - Roberta Marra
- Dipartimento di Medicina Molecolare e Biotecnologie MedicheUniversità degli Studi di Napoli ‘Federico II’ Naples Italy
- CEINGE, Biotecnologie Avanzate Naples Italy
| | - Antonella Gambale
- Dipartimento di Medicina Molecolare e Biotecnologie MedicheUniversità degli Studi di Napoli ‘Federico II’ Naples Italy
- CEINGE, Biotecnologie Avanzate Naples Italy
| | - Domenico Girelli
- Section of Internal Medicine, Department of MedicineUniversity of Verona Verona Italy
| | - Roberta Russo
- Dipartimento di Medicina Molecolare e Biotecnologie MedicheUniversità degli Studi di Napoli ‘Federico II’ Naples Italy
- CEINGE, Biotecnologie Avanzate Naples Italy
| | - Achille Iolascon
- Dipartimento di Medicina Molecolare e Biotecnologie MedicheUniversità degli Studi di Napoli ‘Federico II’ Naples Italy
- CEINGE, Biotecnologie Avanzate Naples Italy
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Mehta N, Zhang D, Li R, Wang T, Gava A, Parthasarathy P, Gao B, Krepinsky JC. Caveolin-1 regulation of Sp1 controls production of the antifibrotic protein follistatin in kidney mesangial cells. Cell Commun Signal 2019; 17:37. [PMID: 30995923 PMCID: PMC6472091 DOI: 10.1186/s12964-019-0351-5] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/08/2019] [Accepted: 04/03/2019] [Indexed: 12/27/2022] Open
Abstract
BACKGROUND We previously showed that caveolin-1 (cav-1), an integral membrane protein, is required for the synthesis of matrix proteins by glomerular mesangial cells (MC). In a previous study to understand how cav-1 is involved in regulating matrix production, we had identified significant upregulation of the antifibrotic protein follistatin in cav-1 knockout MC. Follistatin inhibits the profibrotic effects of several members of the transforming growth factor beta superfamily, in particular the activins. Here, we characterize the molecular mechanism through which cav-1 regulates the expression of follistatin. METHODS Kidneys from cav-1 wild type and knockout (KO) mice were analyzed and primary cultures of MC from cav-1 wild-type and KO mice were utilized. FST promoter deletion constructs were generated to determine the region of the promoter important for mediating FST upregulation in cav-1 KO MC. siRNA-mediated down-regulation and overexpression of Sp1 in conjunction with luciferase activity assays, immunoprecipitation, western blotting and ChiP was used to assess the role of Sp1 in transcriptionally regulating FST expression. Pharmacologic kinase inhibitors and specific siRNA were used to determine the post-translational mechanism through which cav-1 affects Sp1 activity. RESULTS Our results establish that follistatin upregulation occurs at the transcript level. We identified Sp1 as the critical transcription factor regulating activation of the FST promoter in cav-1 KO MC through binding to a region within 123 bp of the transcription start site. We further determined that the lack of cav-1 increases Sp1 nuclear levels and transcriptional activity. This occurred through increased phosphoinositide 3-kinase (PI3K) activity and downstream protein kinase C (PKC) zeta-mediated phosphorylation and activation of Sp1. CONCLUSIONS These findings shed light on the transcriptional mechanism by which cav-1 represses the expression of a major antifibrotic protein, and can inform the development of novel antifibrotic treatment strategies.
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Affiliation(s)
- Neel Mehta
- Division of Nephrology, Department of Medicine, McMaster University, Hamilton, Canada
| | - Dan Zhang
- Division of Nephrology, Department of Medicine, McMaster University, Hamilton, Canada
| | - Renzhong Li
- Division of Nephrology, Department of Medicine, McMaster University, Hamilton, Canada
| | - Tony Wang
- Division of Nephrology, Department of Medicine, McMaster University, Hamilton, Canada
| | - Agata Gava
- Physiological Sciences Graduate Program, Health Sciences Centre, Federal University of Espirito Santo, Vitoria, Brazil
| | | | - Bo Gao
- Division of Nephrology, Department of Medicine, McMaster University, Hamilton, Canada
| | - Joan C Krepinsky
- Division of Nephrology, Department of Medicine, McMaster University, Hamilton, Canada. .,St. Joseph's Hospital, 50 Charlton Ave East, Rm T3311, Hamilton, ON, L8N 4A6, Canada.
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Soleimani A, Pashirzad M, Avan A, Ferns GA, Khazaei M, Hassanian SM. Role of the transforming growth factor-β signaling pathway in the pathogenesis of colorectal cancer. J Cell Biochem 2018; 120:8899-8907. [PMID: 30556274 DOI: 10.1002/jcb.28331] [Citation(s) in RCA: 29] [Impact Index Per Article: 4.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/06/2018] [Accepted: 11/28/2018] [Indexed: 12/22/2022]
Abstract
The transforming growth factor-β (TGF-β) signaling pathway plays an important role in cancer cell proliferation, growth, metastasis, and apoptosis. It has been shown that TGF-β acts as a tumor suppressor in the early stages of the disease, and as a tumor promoter in its late stages. Mutations in the TGF-β signaling components, the TGF-β receptors and cytoplasmic signaling transducers, are frequently observed in colorectal carcinomas. Exploiting specific TGF-β receptor agonist and antagonist with antitumor properties may be a way of controlling cancer progression. This review summarizes the regulatory role of TGF-β signaling in the pathogenesis of colorectal cancer.
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Affiliation(s)
- Atena Soleimani
- Department of Clinical Biochemistry, Faculty of Medicine, Mashhad University of Medical Sciences, Mashhad, Iran.,Student Research Committee, Faculty of Medicine, Mashhad University of Medical Sciences, Mashhad, Iran
| | - Mehran Pashirzad
- Department of Clinical Biochemistry, Faculty of Medicine, Mashhad University of Medical Sciences, Mashhad, Iran
| | - Amir Avan
- Metabolic Syndrome Research Center, Mashhad University of Medical Sciences, Mashhad, Iran.,Department of Modern Sciences and Technologies, Faculty of Medicine, Mashhad University of Medical Sciences, Mashhad, Iran
| | - Gordon A Ferns
- Division of Medical Education, Brighton & Sussex Medical School, Brighton, UK
| | - Majid Khazaei
- Metabolic Syndrome Research Center, Mashhad University of Medical Sciences, Mashhad, Iran.,Department of Medical Physiology, Faculty of Medicine, Mashhad University of Medical Sciences, Mashhad, Iran
| | - Seyed Mahdi Hassanian
- Department of Clinical Biochemistry, Faculty of Medicine, Mashhad University of Medical Sciences, Mashhad, Iran.,Metabolic Syndrome Research Center, Mashhad University of Medical Sciences, Mashhad, Iran
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14
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Qin Z, Xia W, Fisher GJ, Voorhees JJ, Quan T. YAP/TAZ regulates TGF-β/Smad3 signaling by induction of Smad7 via AP-1 in human skin dermal fibroblasts. Cell Commun Signal 2018; 16:18. [PMID: 29695252 PMCID: PMC5918965 DOI: 10.1186/s12964-018-0232-3] [Citation(s) in RCA: 99] [Impact Index Per Article: 14.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/17/2017] [Accepted: 04/17/2018] [Indexed: 12/22/2022] Open
Abstract
Background Transcription factors YAP and TAZ function as the primary mediators of the Hippo pathway. Yet, crosstalk of YAP and TAZ with other signaling pathways remains relatively unexplored. We have explored the impact of YAP and TAZ levels on the TGF-β/Smad signaling pathway in human skin dermal fibroblasts. Methods YAP and TAZ levels in dermal fibroblasts were reduced in dermal fibroblasts by siRNA-mediated knockdown. The effects of YAP and TAZ reduction on TGF-β/Smad signaling were examined by quantitative real-time PCR, Western analysis, and immunostaining. Luciferase reporter assays and electrophoretic mobility shift assays were conducted to investigate the transcription factor DNA-binding and transcriptional activities. Results Knockdown of both YAP and TAZ (YAP/TAZ), but not either separately, impaired TGF-β1-induced Smad3 phosphorylation and Smad3 transcriptional activity, thereby inhibiting the expression of TGF-β target genes. This reduction by reduced levels of YAP/TAZ results from induction of inhibitory Smad7, which inhibits Smad3 phosphorylation and activity by TGF-β1. Conversely, prevention of Smad7 induction restores Smad3 phosphorylation and Smad3 transcriptional activity in fibroblasts that have reduced YAP/TAZ. In agreement with these findings, inhibition of YAP/TAZ transcriptional activity, similar to the reduction of YAP/TAZ levels, also significantly induced Smad7 and impaired TGF-β/Smad signaling. Further investigations revealed that reduced levels of YAP/TAZ led to induction of activator protein-1 (AP-1) activity, Activated AP-1 bound to DNA sequences in the Smad7 gene promoter, and deletion of these AP-1 binding sequences substantially reduced Smad7 promoter reporter activity. Conclusion YAP/TAZ functions in concert with transcription factor AP-1 and Smad7 to regulate TGF-β signaling, in human dermal fibroblasts. Reduction of YAP/TAZ levels leads to activation of AP-1 activity, which induces Smad7. Smad7 suppresses the TGF-β pathway. Electronic supplementary material The online version of this article (10.1186/s12964-018-0232-3) contains supplementary material, which is available to authorized users.
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Affiliation(s)
- Zhaoping Qin
- Department of Dermatology, University of Michigan Medical School, 1301 Catherine, Medical Science I, Room 6447, Ann Arbor, MI, 48109-0609, USA
| | - Wei Xia
- Department of Dermatology, University of Michigan Medical School, 1301 Catherine, Medical Science I, Room 6447, Ann Arbor, MI, 48109-0609, USA
| | - Gary J Fisher
- Department of Dermatology, University of Michigan Medical School, 1301 Catherine, Medical Science I, Room 6447, Ann Arbor, MI, 48109-0609, USA
| | - John J Voorhees
- Department of Dermatology, University of Michigan Medical School, 1301 Catherine, Medical Science I, Room 6447, Ann Arbor, MI, 48109-0609, USA
| | - Taihao Quan
- Department of Dermatology, University of Michigan Medical School, 1301 Catherine, Medical Science I, Room 6447, Ann Arbor, MI, 48109-0609, USA.
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15
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Yan X, Xiong X, Chen YG. Feedback regulation of TGF-β signaling. Acta Biochim Biophys Sin (Shanghai) 2018; 50:37-50. [PMID: 29228156 DOI: 10.1093/abbs/gmx129] [Citation(s) in RCA: 85] [Impact Index Per Article: 12.1] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/31/2017] [Indexed: 12/20/2022] Open
Abstract
Transforming growth factor beta (TGF-β) is a multi-functional polypeptide that plays a critical role in regulating a broad range of cellular functions and physiological processes. Signaling is initiated when TGF-β ligands bind to two types of cell membrane receptors with intrinsic Ser/Thr kinase activity and transmitted by the intracellular Smad proteins, which act as transcription factors to regulate gene expression in the nucleus. Although it is relatively simple and straight-forward, this TGF-β/Smad pathway is regulated by various feedback loops at different levels, including the ligand, the receptor, Smads and transcription, and is thus fine-tuned in terms of signaling robustness, duration, specificity, and plasticity. The precise control gives rise to versatile and context-dependent pathophysiological functions. In this review, we firstly give an overview of TGF-β signaling, and then discuss how each step of TGF-β signaling is finely controlled by distinct modes of feedback mechanisms, involving both protein regulators and miRNAs.
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Affiliation(s)
- Xiaohua Yan
- Department of Biochemistry and Molecular Biology, School of Basic Medical Sciences, Nanchang University, Nanchang 330006, China
| | - Xiangyang Xiong
- Department of Biochemistry and Molecular Biology, School of Basic Medical Sciences, Nanchang University, Nanchang 330006, China
| | - Ye-Guang Chen
- The State Key Laboratory of Membrane Biology, Tsinghua-Peking Center for Life Sciences, School of Life Sciences, Tsinghua University, Beijing 100084, China
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16
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Deng L, Huang L, Guo Q, Shi X, Xu K. CREB1 and Smad3 mediate TGF‑β3‑induced Smad7 expression in rat hepatic stellate cells. Mol Med Rep 2017; 16:8455-8462. [PMID: 28983617 DOI: 10.3892/mmr.2017.7654] [Citation(s) in RCA: 10] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/10/2016] [Accepted: 02/01/2017] [Indexed: 11/05/2022] Open
Abstract
Transforming growth factor (TGF)‑β3 has previously been reported to antagonize hepatic fibrosis in vivo and in vitro. The present study aimed to investigate the mechanism underlying the involvement of TGF‑β3 in hepatic fibrosis. Short hairpin (sh)RNA‑cAMP-responsive element binding protein (CREB) 1 and small interfering (si)RNA‑Smad3 were utilized to silence the expression of CREB1 and Smad3 in hepatic stellate cells (HSCs), whereas the vector pRSV‑CREB1 was used to induce CREB1 overexpression in HSCs. Cells were treated with or without exogenous TGF‑β3 or TGF‑β1, and mRNA and protein expression levels were assessed using reverse transcription‑quantitative polymerase chain reaction and western blot analysis. Untreated cells served as the control group. Exogenous TGF‑β3 increased Smad7 mRNA and protein expression levels in rat HSCs, and CREB1 and Smad3 appeared to be implicated in the mechanism of Smad7. CREB1 knockdown inhibited the TGF‑β3‑induced upregulation of Smad7, whereas its overexpression potentiated the Smad7 upregulation in HSCs; conversely, CREB1 manipulations had no effect on Smad7 expression under basal conditions. In addition, TGF‑β3‑induced Smad7 upregulation was blocked when the activity of p38, a kinase upstream of CREB1, was inhibited. Furthermore, silencing Smad3 resulted in decreased Smad7 expression under basal conditions and in TGF‑β3‑stimulated cells. Notably, Smad7 expression appeared to also be induced by exogenous TGF‑β1, independent of CREB1. The present study demonstrated that TGF‑β3 increased Smad7 expression in HSCs, whereas CREB1 and Smad3 appeared to participate in the mechanism of induction. Smad3 is the key regulator whereas CREB‑1 acts as a co‑regulator. These results suggested that this mechanism may underlie the antagonizing effects of TGF‑β3 on hepatic fibrosis.
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Affiliation(s)
- Liang Deng
- Department of Gastroenterology, The First Affiliated Hospital of Chongqing Medical University, Chongqing 400016, P.R. China
| | - Lu Huang
- Department of Immunology, Children's Hospital of Chongqing Medical University, Chongqing 400014, P.R. China
| | - Qiongya Guo
- Department of Gastroenterology, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei 430022, P.R. China
| | - Xiaoyu Shi
- Department of Gastroenterology, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei 430022, P.R. China
| | - Keshu Xu
- Department of Gastroenterology, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei 430022, P.R. China
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17
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van Caam A, Madej W, Garcia de Vinuesa A, Goumans MJ, Ten Dijke P, Blaney Davidson E, van der Kraan P. TGFβ1-induced SMAD2/3 and SMAD1/5 phosphorylation are both ALK5-kinase-dependent in primary chondrocytes and mediated by TAK1 kinase activity. Arthritis Res Ther 2017; 19:112. [PMID: 28569204 PMCID: PMC5452635 DOI: 10.1186/s13075-017-1302-4] [Citation(s) in RCA: 50] [Impact Index Per Article: 6.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/18/2016] [Accepted: 05/02/2017] [Indexed: 11/26/2022] Open
Abstract
Background Dysregulated transforming growth factor β (TGFβ) signaling is implicated in osteoarthritis development, making normalizing TGFβ signaling a possible therapy. Theoretically, this can be achieved with small molecule inhibitors specifically targeting the various TGFβ receptors and downstream mediators. In this study we explore in primary chondrocytes the use of small molecule inhibitors to target TGFβ-induced pSmad1/5/9-, pSmad2/3- and TGFβ-activated kinase 1 (TAK1)-dependent signaling. Method Primary bovine chondrocytes and explants were isolated from metacarpophalangeal joints. To modulate TGFβ signaling the activin receptor-like kinase (ALK)1/2/3/6 inhibitor LDN-193189, the ALK4/5/7 inhibitor SB-505124 and the TAK1 inhibitor (5Z)-7-Oxozeaenol were used. pSmad1/5 and pSmad2 were measured using western blot analysis and TGFβ1-induced gene expression was measured using quantitative real time PCR (qPCR). Results In chondrocytes, TGFβ1 strongly induced both pSmad1/5 and pSmad2. Remarkably, LDN-193189 did not inhibit TGFβ-induced pSmad1/5. In contrast, SB-505124 did inhibit both TGFβ-induced Smad2 and Smad1/5 phosphorylation. Furthermore, (5Z)-7-Oxozeaenol also profoundly inhibited TGFβ-induced pSmad2 and pSmad1/5. Importantly, both SB-505124 and (5Z)-7-Oxozeaenol did not significantly inhibit constitutively active ALK1, making an off-target effect unlikely. Additionally, LDN-193189 was able to potently inhibit BMP2/7/9-induced pSmad1/5, showing its functionality. On gene expression, LDN-193189 did not affect TGFβ1-induced regulation, whereas both SB-505124 and (5Z)-7-Oxozeaenol did. Similar results were obtained in cartilage explants, although pSmad1/5 was not strongly induced by addition of TGFβ1. Conclusion Our data suggest that ALK5 kinase activity plays a central role in both TGFβ-induced Smad1/5 and Smad2/3 phosphorylation, making it difficult to separate both pathways with the use of currently available small molecule inhibitors. Furthermore, our data regarding (5Z)-7-Oxozeaenol suggest that TAK1 facilitates Smad-dependent signaling. Electronic supplementary material The online version of this article (doi:10.1186/s13075-017-1302-4) contains supplementary material, which is available to authorized users.
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Affiliation(s)
- Arjan van Caam
- Experimental Rheumatology, Radboud University Medical Center, Nijmegen, The Netherlands.
| | - Wojciech Madej
- Experimental Rheumatology, Radboud University Medical Center, Nijmegen, The Netherlands.,Orthopaedics Research Lab, Radboud University Medical Center, Nijmegen, The Netherlands
| | - Amaya Garcia de Vinuesa
- Department of Molecular Cell Biology and Cancer Genomics Centre Netherlands, Leiden University Medical Center, Leiden, The Netherlands
| | - Marie-José Goumans
- Department of Molecular Cell Biology and Cancer Genomics Centre Netherlands, Leiden University Medical Center, Leiden, The Netherlands
| | - Peter Ten Dijke
- Department of Molecular Cell Biology and Cancer Genomics Centre Netherlands, Leiden University Medical Center, Leiden, The Netherlands
| | | | - Peter van der Kraan
- Experimental Rheumatology, Radboud University Medical Center, Nijmegen, The Netherlands
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Abstract
Inhibitory Smads (I-Smads) have conserved carboxy-terminal MH2 domains but highly divergent amino-terminal regions when compared with receptor-regulated Smads (R-Smads) and common-partner Smads (co-Smads). Smad6 preferentially inhibits Smad signaling initiated by the bone morphogenetic protein (BMP) type I receptors ALK-3 and ALK-6, whereas Smad7 inhibits both transforming growth factor β (TGF-β)- and BMP-induced Smad signaling. I-Smads also regulate some non-Smad signaling pathways. Here, we discuss the vertebrate I-Smads, their roles as inhibitors of Smad activation and regulators of receptor stability, as scaffolds for non-Smad signaling, and their possible roles in the nucleus. We also discuss the posttranslational modification of I-Smads, including phosphorylation, ubiquitylation, acetylation, and methylation.
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Affiliation(s)
- Keiji Miyazawa
- Department of Biochemistry, Interdisciplinary Graduate School of Medicine, University of Yamanashi, Chuo, Yamanashi 409-3898, Japan
| | - Kohei Miyazono
- Department of Molecular Pathology, Graduate School of Medicine, The University of Tokyo, Bunkyo-ku, Tokyo 113-0033, Japan
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19
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TIEG1 Represses Smad7-Mediated Activation of TGF-β1/Smad Signaling in Keloid Pathogenesis. J Invest Dermatol 2017; 137:1051-1059. [PMID: 28108300 DOI: 10.1016/j.jid.2016.12.019] [Citation(s) in RCA: 37] [Impact Index Per Article: 4.6] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/07/2016] [Revised: 12/14/2016] [Accepted: 12/19/2016] [Indexed: 01/16/2023]
Abstract
Transforming growth factor-β (TGF-β)/Smad signaling plays a key role in excessive fibrosis and keloid formations. Smad7 is a negative feedback regulator that prevents activation of TGF-β/Smad signaling. However, the regulatory mechanism for Smad7 in the keloid pathogenic process remains elusive. Here, we show that expression of TIEG1 is markedly higher in keloid fibroblasts, whereas protein, mRNA, and promoter activity levels of Smad7 are decreased. When TIEG1 was knocked down with small interfering RNA, both the promoter activity and protein expression of Smad7 were increased, whereas collagen production and the proliferation, migration, and invasion of keloid fibroblasts were decreased. In contrast, TIEG1 overexpression led to a decrease in Smad7 expression and Smad7 promoter activity. Upon TGF-β1 stimulation, TIEG1 promoted Smad2 phosphorylation by down-regulating Smad7. Luciferase reporter assays and chromatin immunoprecipitation assays further showed that TIEG1 can directly bind a GC-box/Sp1 site located between nucleotides -1392 and -1382 in the Smad7 promoter to repress Smad7 promoter activity. Taken together, these findings show that TIEG1 is highly expressed in human keloids and that it directly binds and represses Smad7 promoter-mediated activation of TGF-β/Smad2 signaling, thus providing clues for development of TIEG1 blocking strategies for therapy or prophylaxis of keloids.
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20
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Abstract
Cytokines of the transforming growth factor β (TGF-β) family, including TGF-βs, bone morphogenic proteins (BMPs), activins, and Nodal, play crucial roles in embryonic development and adult tissue homeostasis by regulating cell proliferation, survival, and differentiation, as well as stem-cell self-renewal and lineage-specific differentiation. Smad proteins are critical downstream mediators of these signaling activities. In addition to regulating the transcription of direct target genes of TGF-β, BMP, activin, or Nodal, Smad proteins also participate in extensive cross talk with other signaling pathways, often in a cell-type- or developmental stage-specific manner. These combinatorial signals often produce context-, time-, and location-dependent biological outcomes that are critical for development. This review discusses recent progress in our understanding of the cross talk between Smad proteins and signaling pathways of Wnt, Notch, Hippo, Hedgehog (Hh), mitogen-activated protein (MAP), kinase, phosphoinositide 3-kinase (PI3K)-Akt, nuclear factor κB (NF-κB), and Janus kinase/signal transducers and activators of transcription (JAK/STAT) pathways.
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Affiliation(s)
- Kunxin Luo
- Department of Molecular and Cell Biology, University of California, Berkeley, and Life Sciences Division, Lawrence Berkeley National Laboratory, Berkeley, California 94720
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21
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Luo K. Signaling Cross Talk between TGF-β/Smad and Other Signaling Pathways. Cold Spring Harb Perspect Biol 2017. [PMID: 27836834 DOI: 10.1101/cshperspect] [Citation(s) in RCA: 33] [Impact Index Per Article: 4.1] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/14/2022]
Abstract
Cytokines of the transforming growth factor β (TGF-β) family, including TGF-βs, bone morphogenic proteins (BMPs), activins, and Nodal, play crucial roles in embryonic development and adult tissue homeostasis by regulating cell proliferation, survival, and differentiation, as well as stem-cell self-renewal and lineage-specific differentiation. Smad proteins are critical downstream mediators of these signaling activities. In addition to regulating the transcription of direct target genes of TGF-β, BMP, activin, or Nodal, Smad proteins also participate in extensive cross talk with other signaling pathways, often in a cell-type- or developmental stage-specific manner. These combinatorial signals often produce context-, time-, and location-dependent biological outcomes that are critical for development. This review discusses recent progress in our understanding of the cross talk between Smad proteins and signaling pathways of Wnt, Notch, Hippo, Hedgehog (Hh), mitogen-activated protein (MAP), kinase, phosphoinositide 3-kinase (PI3K)-Akt, nuclear factor κB (NF-κB), and Janus kinase/signal transducers and activators of transcription (JAK/STAT) pathways.
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Affiliation(s)
- Kunxin Luo
- Department of Molecular and Cell Biology, University of California, Berkeley, and Life Sciences Division, Lawrence Berkeley National Laboratory, Berkeley, California 94720
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22
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Yu J, Lei R, Zhuang X, Li X, Li G, Lev S, Segura MF, Zhang X, Hu G. MicroRNA-182 targets SMAD7 to potentiate TGFβ-induced epithelial-mesenchymal transition and metastasis of cancer cells. Nat Commun 2016; 7:13884. [PMID: 27996004 PMCID: PMC5187443 DOI: 10.1038/ncomms13884] [Citation(s) in RCA: 100] [Impact Index Per Article: 11.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/02/2015] [Accepted: 11/09/2016] [Indexed: 12/13/2022] Open
Abstract
The transforming growth factor β (TGFβ) pathway plays critical roles during cancer cell epithelial-mesenchymal transition (EMT) and metastasis. SMAD7 is both a transcriptional target and a negative regulator of TGFβ signalling, thus mediating a negative feedback loop that may potentially restrain TGFβ responses of cancer cells. Here, however, we show that TGFβ treatment induces SMAD7 transcription but not its protein level in a panel of cancer cells. Mechanistic studies reveal that TGFβ activates the expression of microRNA-182 (miR-182), which suppresses SMAD7 protein. miR-182 silencing leads to SMAD7 upregulation on TGFβ treatment and prevents TGFβ-induced EMT and invasion of cancer cells. Overexpression of miR-182 promotes breast tumour invasion and TGFβ-induced osteoclastogenesis for bone metastasis. Furthermore, miR-182 expression inversely correlates with SMAD7 protein in human tumour samples. Therefore, our data reveal the miR-182-mediated disruption of TGFβ self-restraint and provide a mechanism to explain the unleashed TGFβ responses in metastatic cancer cells. SMAD7 is a transcriptional target and a negative regulator of TGFβ signalling forming a negative feedback loop. Here the authors show that in cancer cells TGFβ activates the expression of microRNA-182 that suppresses SMAD7 protein, promoting TGFβ-mediated breast tumour invasion and bone metastasis.
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Affiliation(s)
- Jingyi Yu
- The Key Laboratory of Stem Cell Biology, Institute of Health Sciences, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences &Shanghai Jiao Tong University School of Medicine, University of Chinese Academy of Sciences, Shanghai 200031, China
| | - Rong Lei
- The Key Laboratory of Stem Cell Biology, Institute of Health Sciences, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences &Shanghai Jiao Tong University School of Medicine, University of Chinese Academy of Sciences, Shanghai 200031, China
| | - Xueqian Zhuang
- The Key Laboratory of Stem Cell Biology, Institute of Health Sciences, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences &Shanghai Jiao Tong University School of Medicine, University of Chinese Academy of Sciences, Shanghai 200031, China
| | - Xiaoxun Li
- The Key Laboratory of Stem Cell Biology, Institute of Health Sciences, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences &Shanghai Jiao Tong University School of Medicine, University of Chinese Academy of Sciences, Shanghai 200031, China
| | - Gang Li
- The Key Laboratory of Stem Cell Biology, Institute of Health Sciences, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences &Shanghai Jiao Tong University School of Medicine, University of Chinese Academy of Sciences, Shanghai 200031, China
| | - Sima Lev
- Molecular Cell Biology Department, Weizmann Institute of Science, Rehovot 76100, Israel
| | - Miguel F Segura
- Department of Pathology, New York University School of Medicine, New York, New York 10016, USA
| | - Xue Zhang
- The Key Laboratory of Stem Cell Biology, Institute of Health Sciences, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences &Shanghai Jiao Tong University School of Medicine, University of Chinese Academy of Sciences, Shanghai 200031, China
| | - Guohong Hu
- The Key Laboratory of Stem Cell Biology, Institute of Health Sciences, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences &Shanghai Jiao Tong University School of Medicine, University of Chinese Academy of Sciences, Shanghai 200031, China
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23
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Madej W, van Caam A, Blaney Davidson E, Buma P, van der Kraan PM. Unloading results in rapid loss of TGFβ signaling in articular cartilage: role of loading-induced TGFβ signaling in maintenance of articular chondrocyte phenotype? Osteoarthritis Cartilage 2016; 24:1807-1815. [PMID: 27268943 DOI: 10.1016/j.joca.2016.05.018] [Citation(s) in RCA: 28] [Impact Index Per Article: 3.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 12/22/2015] [Revised: 04/28/2016] [Accepted: 05/23/2016] [Indexed: 02/02/2023]
Abstract
OBJECTIVE Recently it was shown that loading of articular cartilage explants activates TGFβ signaling. Here we investigated if in vivo chondrocytes express permanently high TGFβ signaling, and the consequence of the loss of compressive loading-mediated TGFβ signaling on chondrocyte function and phenotype. METHOD Bovine articular cartilage explants were collected within 10 min post mortem and stained immediately and after 30, 60 and 360 min for phosphorylated-Smad2, indicating active TGFβ signaling. Explants were unloaded for 48 h and subsequently repeatedly loaded with a compressive load of 3 MPa. In addition, explants were cultured unloaded for 2 weeks and the effect of loading or exogenous TGFβ on proteoglycan level and chondrocyte phenotype (Col10a1 mRNA expression) was analyzed. RESULTS Unloading of articular cartilage results in rapid loss of TGFβ signaling while subsequent compressive loading swiftly restored this. Loading and exogenous TGFβ enhanced expression of TGFβ1 and ALK5. Unloading of explants for 2 weeks resulted in proteoglycan loss and increased Col10a1 expression. Both loading and exogenous TGFβ inhibited elevated Col10a1 expression but not proteoglycan loss. CONCLUSION Our data might imply that in vivo regular physiological loading of articular cartilage leads to enduring TGFβ signaling and TGFβ-induced gene expression. We propose a hypothetical model in which loading activates a self-perpetuating system that prevents hypertrophic differentiation of chondrocytes and is crucial for cartilage homeostasis.
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Affiliation(s)
- W Madej
- Orthopedic Research Laboratory, Radboudumc, PO Box 9101, 6500 HB Nijmegen, The Netherlands
| | - A van Caam
- Experimental Rheumatology, Radboudumc, PO Box 9101, 6500 HB Nijmegen, The Netherlands
| | - E Blaney Davidson
- Experimental Rheumatology, Radboudumc, PO Box 9101, 6500 HB Nijmegen, The Netherlands
| | - P Buma
- Orthopedic Research Laboratory, Radboudumc, PO Box 9101, 6500 HB Nijmegen, The Netherlands
| | - P M van der Kraan
- Experimental Rheumatology, Radboudumc, PO Box 9101, 6500 HB Nijmegen, The Netherlands.
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van Caam A, Madej W, Thijssen E, Garcia de Vinuesa A, van den Berg W, Goumans MJ, Ten Dijke P, Blaney Davidson E, van der Kraan PM. Expression of TGFβ-family signalling components in ageing cartilage: age-related loss of TGFβ and BMP receptors. Osteoarthritis Cartilage 2016; 24:1235-45. [PMID: 26975812 DOI: 10.1016/j.joca.2016.02.008] [Citation(s) in RCA: 34] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 09/10/2015] [Revised: 02/11/2016] [Accepted: 02/26/2016] [Indexed: 02/07/2023]
Abstract
OBJECTIVE Ageing is the main risk factor for osteoarthritis (OA). We investigated if expression of transforming growth factor β (TGFβ)-family components, a family which is crucial for the maintenance of healthy articular cartilage, is altered during ageing in cartilage. Moreover, we investigated the functional significance of selected age-related changes. DESIGN Age-related changes in expression of TGFβ-family members were analysed by quantitative PCR in healthy articular cartilage obtained from 42 cows (age: ¾-10 years). To obtain functional insight of selected changes, cartilage explants were stimulated with TGFβ1 or bone morphogenetic protein (BMP) 9, and TGFβ1 and BMP response genes were measured. RESULTS Age-related cartilage thinning and loss of collagen type 2a1 expression (∼256-fold) was observed, validating our data set for studying ageing in cartilage. Expression of the TGFβ-family type I receptors; bAlk2, bAlk3, bAlk4 and bAlk5 dropped significantly with advancing age, whereas bAlk1 expression did not. Of the type II receptors, expression of bBmpr2 decreased significantly. Type III receptor expression was unaffected by ageing. Expression of the ligands bTgfb1 and bGdf5 also decreased with age. In explants, an age-related decrease in TGFβ1-response was observed for the pSmad3-dependent gene bSerpine1 (P = 0.016). In contrast, ageing did not affect BMP9 signalling, an Alk1 ligand, as measured by expression of the pSmad1/5 dependent gene bId1. CONCLUSIONS Ageing negatively affects both the TGFβ-ALK5 and BMP-BMPR signalling routes, and aged chondrocytes display a lowered pSmad3-dependent response to TGFβ1. Because pSmad3 signalling is essential for cartilage homeostasis, we propose that this change contributes to OA development.
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Affiliation(s)
- A van Caam
- Experimental Rheumatology, Radboud University Medical Center, Nijmegen, The Netherlands
| | - W Madej
- Experimental Rheumatology, Radboud University Medical Center, Nijmegen, The Netherlands; Orthopaedics Research Lab, Radboud University Medical Center, Nijmegen, The Netherlands
| | - E Thijssen
- Experimental Rheumatology, Radboud University Medical Center, Nijmegen, The Netherlands
| | - A Garcia de Vinuesa
- Department of Molecular Cell Biology and Cancer Genomics Centre Netherlands, Leiden University Medical Center, Leiden, The Netherlands
| | - W van den Berg
- Experimental Rheumatology, Radboud University Medical Center, Nijmegen, The Netherlands
| | - M-J Goumans
- Department of Molecular Cell Biology and Cancer Genomics Centre Netherlands, Leiden University Medical Center, Leiden, The Netherlands
| | - P Ten Dijke
- Department of Molecular Cell Biology and Cancer Genomics Centre Netherlands, Leiden University Medical Center, Leiden, The Netherlands
| | - E Blaney Davidson
- Experimental Rheumatology, Radboud University Medical Center, Nijmegen, The Netherlands
| | - P M van der Kraan
- Experimental Rheumatology, Radboud University Medical Center, Nijmegen, The Netherlands.
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The high affinity ALK1-ligand BMP9 induces a hypertrophy-like state in chondrocytes that is antagonized by TGFβ1. Osteoarthritis Cartilage 2015; 23:985-95. [PMID: 25681563 DOI: 10.1016/j.joca.2015.02.007] [Citation(s) in RCA: 25] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 09/01/2014] [Revised: 01/26/2015] [Accepted: 02/03/2015] [Indexed: 02/02/2023]
Abstract
OBJECTIVE In osteoarthritic cartilage, expression of the receptor ALK1 correlates with markers of deleterious chondrocyte hypertrophy. Recently, bone morphogenetic protein 9 (BMP9) was identified as a high affinity ligand for ALK1. Therefore, we studied if BMP9 signaling results in expression of hypertrophy markers in chondrocytes. Furthermore, because transforming growth factorß1 (TGFβ1) is a well known anti-hypertrophic factor, the interaction between BMP9 and TGFβ1 signaling was also studied. DESIGN Primary chondrocytes were isolated from bovine cartilage and stimulated with BMP9 and/or TGFβ1 to measure intracellular signaling via pSmads with the use of Western blot. Expression of Smad-responsive genes or hypertrophy-marker genes was measured using qPCR. To confirm observations on TGFβ/Smad3 responsive genes, a Smad3-dependent CAGA12-luc transcriptional reporter assay was performed in the chondrocyte G6 cell line. RESULTS In primary chondrocytes, BMP9 potently induced phosphorylation of Smad1/5 and Smad2 to a lesser extent. BMP9-induced Smad1/5 phosphorylation was rapidly (2 h) reflected in gene expression, whereas Smad2 phosphorylation was not. Remarkably, BMP9 and TGFβ1 dose-dependently synergized on Smad2 phosphorylation, and showed an additive effect on expression of Smad3-dependent genes like bSerpine1 after 24 h. The activation of the TGFβ/Smad3 signaling cascade was confirmed using the CAGA12-luc transcriptional reporter. BMP9 selectively induced bAlpl and bColX expression, which are considered early markers of cellular hypertrophy, but this was potently antagonized by addition of a low dose of TGFβ1. CONCLUSIONS This study shows that in vitro in chondrocytes, BMP9 potently induces pSmad1/5 and a chondrocyte hypertrophy-like state, which is potently blocked by TGFβ1. This observation underlines the importance of TGFβ1 in maintenance of chondrocyte phenotype.
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Fortini BK, Tring S, Plummer SJ, Edlund CK, Moreno V, Bresalier RS, Barry EL, Church TR, Figueiredo JC, Casey G. Multiple functional risk variants in a SMAD7 enhancer implicate a colorectal cancer risk haplotype. PLoS One 2014; 9:e111914. [PMID: 25375357 PMCID: PMC4223076 DOI: 10.1371/journal.pone.0111914] [Citation(s) in RCA: 26] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/16/2014] [Accepted: 10/01/2014] [Indexed: 12/31/2022] Open
Abstract
Genome-wide association studies (GWAS) of colorectal cancer (CRC) have led to the identification of a number of common variants associated with modest risk. Several risk variants map within the vicinity of TGFβ/BMP signaling pathway genes, including rs4939827 within an intron of SMAD7 at 18q21.1. A previous study implicated a novel SNP (novel 1 or rs58920878) as a functional variant within an enhancer element in SMAD7 intron 4. In this study, we show that four SNPs including novel 1 (rs6507874, rs6507875, rs8085824, and rs58920878) in linkage disequilibrium (LD) with the index SNP rs4939827 demonstrate allele-specific enhancer effects in a large, multi-component enhancer of SMAD7. All four SNPs demonstrate allele-specific protein binding to nuclear extracts of CRC cell lines. Furthermore, some of the risk-associated alleles correlate with increased expression of SMAD7 in normal colon tissues. Finally, we show that the enhancer is responsive to BMP4 stimulation. Taken together, we propose that the associated CRC risk at 18q21.1 is due to four functional variants that regulate SMAD7 expression and potentially perturb a BMP negative feedback loop in TGFβ/BMP signaling pathways.
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Affiliation(s)
- Barbara K. Fortini
- Department of Preventive Medicine, University of Southern California, Los Angeles, California, United States of America
- * E-mail:
| | - Stephanie Tring
- Department of Preventive Medicine, University of Southern California, Los Angeles, California, United States of America
| | - Sarah J. Plummer
- Department of Preventive Medicine, University of Southern California, Los Angeles, California, United States of America
| | - Christopher K. Edlund
- Department of Preventive Medicine, University of Southern California, Los Angeles, California, United States of America
| | - Victor Moreno
- Cancer Prevention and Control Program, Catalan Institute of Oncology, CIBERESP and University of Barcelona, Hospitalet de Llobregat, Barcelona, Spain
| | - Robert S. Bresalier
- Department of Gastroenterology, Hepatology and Nutrition, University of Texas MD Anderson Cancer Center, Houston, Texas, United States of America
| | - Elizabeth L. Barry
- Department of Community and Family Medicine, Geisel School of Medicine at Dartmouth, Lebanon, New Hampshire, United States of America
| | - Timothy R. Church
- Division of Environmental Health Sciences, University of Minnesota School of Public Health, Minneapolis, Minnesota, United States of America
| | - Jane C. Figueiredo
- Department of Preventive Medicine, University of Southern California, Los Angeles, California, United States of America
| | - Graham Casey
- Department of Preventive Medicine, University of Southern California, Los Angeles, California, United States of America
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Xiao X, Gaffar I, Guo P, Wiersch J, Fischbach S, Peirish L, Song Z, El-Gohary Y, Prasadan K, Shiota C, Gittes GK. M2 macrophages promote beta-cell proliferation by up-regulation of SMAD7. Proc Natl Acad Sci U S A 2014; 111:E1211-E1220. [PMID: 24639504 PMCID: PMC3977272 DOI: 10.1073/pnas.1321347111] [Citation(s) in RCA: 234] [Impact Index Per Article: 21.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/15/2022] Open
Abstract
Determination of signaling pathways that regulate beta-cell replication is critical for beta-cell therapy. Here, we show that blocking pancreatic macrophage infiltration after pancreatic duct ligation (PDL) completely inhibits beta-cell proliferation. The TGFβ superfamily signaling inhibitor SMAD7 was significantly up-regulated in beta cells after PDL. Beta cells failed to proliferate in response to PDL in beta-cell-specific SMAD7 mutant mice. Forced expression of SMAD7 in beta cells by itself was sufficient to promote beta-cell proliferation in vivo. M2, rather than M1 macrophages, seem to be the inducers of SMAD7-mediated beta-cell proliferation. M2 macrophages not only release TGFβ1 to directly induce up-regulation of SMAD7 in beta cells but also release EGF to activate EGF receptor signaling that inhibits TGFβ1-activated SMAD2 nuclear translocation, resulting in TGFβ signaling inhibition. SMAD7 promotes beta-cell proliferation by increasing CyclinD1 and CyclinD2, and by inducing nuclear exclusion of p27. Our study thus reveals a molecular pathway to potentially increase beta-cell mass through enhanced SMAD7 activity induced by extracellular stimuli.
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Affiliation(s)
| | | | | | - John Wiersch
- Division of Pediatric Surgery, Children’s Hospital of Pittsburgh, University of Pittsburgh School of Medicine, Pittsburgh, PA 15224
| | - Shane Fischbach
- Division of Pediatric Surgery, Children’s Hospital of Pittsburgh, University of Pittsburgh School of Medicine, Pittsburgh, PA 15224
| | - Lauren Peirish
- Division of Pediatric Surgery, Children’s Hospital of Pittsburgh, University of Pittsburgh School of Medicine, Pittsburgh, PA 15224
| | - Zewen Song
- Division of Pediatric Surgery, Children’s Hospital of Pittsburgh, University of Pittsburgh School of Medicine, Pittsburgh, PA 15224
| | - Yousef El-Gohary
- Division of Pediatric Surgery, Children’s Hospital of Pittsburgh, University of Pittsburgh School of Medicine, Pittsburgh, PA 15224
| | - Krishna Prasadan
- Division of Pediatric Surgery, Children’s Hospital of Pittsburgh, University of Pittsburgh School of Medicine, Pittsburgh, PA 15224
| | - Chiyo Shiota
- Division of Pediatric Surgery, Children’s Hospital of Pittsburgh, University of Pittsburgh School of Medicine, Pittsburgh, PA 15224
| | - George K. Gittes
- Division of Pediatric Surgery, Children’s Hospital of Pittsburgh, University of Pittsburgh School of Medicine, Pittsburgh, PA 15224
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Guerquin MJ, Charvet B, Nourissat G, Havis E, Ronsin O, Bonnin MA, Ruggiu M, Olivera-Martinez I, Robert N, Lu Y, Kadler KE, Baumberger T, Doursounian L, Berenbaum F, Duprez D. Transcription factor EGR1 directs tendon differentiation and promotes tendon repair. J Clin Invest 2013; 123:3564-76. [PMID: 23863709 DOI: 10.1172/jci67521] [Citation(s) in RCA: 192] [Impact Index Per Article: 16.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/24/2012] [Accepted: 05/21/2013] [Indexed: 12/27/2022] Open
Abstract
Tendon formation and repair rely on specific combinations of transcription factors, growth factors, and mechanical parameters that regulate the production and spatial organization of type I collagen. Here, we investigated the function of the zinc finger transcription factor EGR1 in tendon formation, healing, and repair using rodent animal models and mesenchymal stem cells (MSCs). Adult tendons of Egr1-/- mice displayed a deficiency in the expression of tendon genes, including Scx, Col1a1, and Col1a2, and were mechanically weaker compared with their WT littermates. EGR1 was recruited to the Col1a1 and Col2a1 promoters in postnatal mouse tendons in vivo. Egr1 was required for the normal gene response following tendon injury in a mouse model of Achilles tendon healing. Forced Egr1 expression programmed MSCs toward the tendon lineage and promoted the formation of in vitro-engineered tendons from MSCs. The application of EGR1-producing MSCs increased the formation of tendon-like tissues in a rat model of Achilles tendon injury. We provide evidence that the ability of EGR1 to promote tendon differentiation is partially mediated by TGF-β2. This study demonstrates EGR1 involvement in adult tendon formation, healing, and repair and identifies Egr1 as a putative target in tendon repair strategies.
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IHG-1 must be localised to mitochondria to decrease Smad7 expression and amplify TGF-β1-induced fibrotic responses. BIOCHIMICA ET BIOPHYSICA ACTA-MOLECULAR CELL RESEARCH 2013; 1833:1969-78. [PMID: 23567938 DOI: 10.1016/j.bbamcr.2013.03.027] [Citation(s) in RCA: 16] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Received: 08/07/2012] [Revised: 02/28/2013] [Accepted: 03/26/2013] [Indexed: 01/15/2023]
Abstract
TGF-β1 is a prototypic profibrotic cytokine and major driver of fibrosis in the kidney and other organs. Induced in high glucose-1 (IHG-1) is a mitochondrial protein which we have recently reported to be associated with renal disease. IHG-1 amplifies responses to TGF-β1 and regulates mitochondrial biogenesis by stabilising the transcriptional co-activator peroxisome proliferator-activated receptor gamma coactivator-1-alpha. Here we report that the mitochondrial localisation of IHG-1 is pivotal in the amplification of TGF-β1 signalling. We demonstrate that IHG-1 expression is associated with repression of the endogenous TGF-β1 inhibitor Smad7. Intriguingly, expression of a non-mitochondrial deletion mutant of IHG-1 (Δmts-IHG-1) repressed TGF-β1 fibrotic signalling in renal epithelial cells. In cells expressing Δmts-IHG-1 fibrotic responses including CCN2/connective tissue growth factor, fibronectin and jagged-1 expression were reduced following stimulation with TGF-β1. Δmts-IHG-1 modulation of TGF-β1 signalling was associated with increased Smad7 protein expression. Δmts-IHG-1 modulated TGF-β1 activity by increasing Smad7 protein expression as it failed to inhibit TGF-β1 transcriptional responses when endogenous Smad7 expression was knocked down. These data indicate that mitochondria modulate TGF-β1 signal transduction and that IHG-1 is a key player in this modulation.
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Zhang TY, Labonté B, Wen XL, Turecki G, Meaney MJ. Epigenetic mechanisms for the early environmental regulation of hippocampal glucocorticoid receptor gene expression in rodents and humans. Neuropsychopharmacology 2013; 38:111-23. [PMID: 22968814 PMCID: PMC3521971 DOI: 10.1038/npp.2012.149] [Citation(s) in RCA: 239] [Impact Index Per Article: 19.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 04/03/2012] [Revised: 05/31/2012] [Accepted: 05/31/2012] [Indexed: 01/22/2023]
Abstract
Parental care influences development across mammals. In humans such influences include effects on phenotypes, such as stress reactivity, which determine individual differences in the vulnerability for affective disorders. Thus, the adult offspring of rat mothers that show an increased frequency of pup licking/grooming (ie, high LG mothers) show increased hippocampal glucocorticoid receptor (GR) expression and more modest hypothalamic-pituitary-adrenal responses to stress compared with the offspring of low LG mothers. In humans, childhood maltreatment associates decreased hippocampal GR expression and increased stress responses in adulthood. We review the evidence suggesting that such effects are mediated by epigenetic mechanisms, including DNA methylation and hydroxymethylation across GR promoter regions. We also present new findings revealing associated histone post-translational modifications of a critical GR promoter in rat hippocampus. Taken together these existing evidences are consistent with the idea that parental influences establish stable phenotypic variation in the offspring through effects on intracellular signaling pathways that regulate the epigenetic state and function of specific regions of the genome.
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Affiliation(s)
- Tie Yuan Zhang
- Sackler Program for Epigenetics Psychobiology and Departments of Psychiatry and Neurology & Neurosurgery, McGill University, Montreal, Quebec, Canada
| | - Benoit Labonté
- Sackler Program for Epigenetics Psychobiology and Departments of Psychiatry and Neurology & Neurosurgery, McGill University, Montreal, Quebec, Canada
| | - Xiang Lan Wen
- Sackler Program for Epigenetics Psychobiology and Departments of Psychiatry and Neurology & Neurosurgery, McGill University, Montreal, Quebec, Canada
| | - Gustavo Turecki
- Sackler Program for Epigenetics Psychobiology and Departments of Psychiatry and Neurology & Neurosurgery, McGill University, Montreal, Quebec, Canada
| | - Michael J Meaney
- Sackler Program for Epigenetics Psychobiology and Departments of Psychiatry and Neurology & Neurosurgery, McGill University, Montreal, Quebec, Canada
- Singapore Institute for Clinical Sciences, Singapore, Singapore
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Smad7 is a transforming growth factor-beta-inducible mediator of apoptosis in granulosa cells. Fertil Steril 2012; 97:1452-9.e1-6. [PMID: 22656308 DOI: 10.1016/j.fertnstert.2012.03.024] [Citation(s) in RCA: 27] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/08/2011] [Revised: 03/13/2012] [Accepted: 03/14/2012] [Indexed: 02/08/2023]
Abstract
OBJECTIVE To determine the functional role of Smad7 in granulosa cells. DESIGN Granulosa cell culture and molecular biological techniques were used to investigate regulation and function of Smad7. SETTING Research laboratory. ANIMAL(S) C57bl/j hybrid mouse. INTERVENTION(S) Primary mouse granulosa cells were isolated and grown in culture for all messenger RNA expression experiments. Smad7 promoter constructs were evaluated with a luciferase reporter system in SIGC cells to determine sites activating Smad7 expression. MAIN OUTCOME MEASURE(S) Overexpression (Smad7 complementary DNA) and downregulation (Smad7 small interfering RNA) of Smad7 in primary mouse granulosa cells were used to evaluate the functional role of Smad7 in granulosa cells. RESULT(S) Smad7 expression was upregulated by treatment with transforming growth factor-β (TGF-β) but not activin or activation of the cyclic adenosine monophosphate pathway. The promoter of Smad7 was activated by TGF-β. Truncation of the promoter or mutation of the Smad response element at -141 eliminated TGF-β activation of the promoter. Smad3 was not specifically required for TGF-β-stimulated expression of Smad7, though activation of the TGFBR1 receptor was. When Smad7 was overexpressed in granulosa cells, apoptosis was markedly increased. When Smad7 expression was reduced with small interfering RNA, then the TGF-β-induced apoptosis was blocked. CONCLUSION(S) Smad7 mediates apoptosis induced by TGF-β in mouse granulosa cells, suggesting that dysregulation of Smad7 could impair folliculogenesis.
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Targeting Stat3 and Smad7 to restore TGF-β cytostatic regulation of tumor cells in vitro and in vivo. Oncogene 2012; 32:2433-41. [PMID: 22751114 PMCID: PMC3655378 DOI: 10.1038/onc.2012.260] [Citation(s) in RCA: 63] [Impact Index Per Article: 4.8] [Reference Citation Analysis] [Abstract] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/18/2023]
Abstract
Transforming Growth Factor-β (TGF-β) and Epidermal Growth Factor (EGF) signaling pathways are both independently implicated as key regulators in tumor formation and progression. Here, we report that the tumor-associated overexpression of epidermal growth factor receptor (EGFR) desensitizes TGF-β signaling and its cytostatic regulation through specific and persistent Stat3 activation and Smad7 induction in vivo. In human tumor cell lines, reduction of TGF-β-mediated Smad2 phosphorylation, nuclear translocation and Smad3 target gene activation were observed when EGFR was overexpressed, but not in cells that expressed EGFR at normal levels. We identified Stat3, which is activated specifically and persistently by overexpressed EGFR, as a key signaling molecule responsible for the reduced TGF-β sensitivity. Stable knockdown of Stat3 using small hairpin RNA(shRNA) in Head and Neck (HN5) and Epidermoid (A431) tumor cell lines resulted in reduced growth compared with control shRNA-transfected cells when grown as subcutaneous tumor xenografts. Furthermore, xenografts with Stat3 knockdown displayed increased Smad3 transcriptional activity, increased Smad2 phosphorylation and decreased Smad7 expression compared with control xenografts in vivo. Consistently, Smad7 mRNA and protein expression was also significantly reduced when EGFR activity was blocked by a specific tyrosine kinase inhibitor, AG1478, or in Stat3 knockdown tumors. Similarly, Smad7 knockdown also resulted in enhanced Smad3 transcriptional activity in vivo. Importantly, there was no uptake of subcutaneous HN5 xenografts with Smad7 knockdown. Taken together, we demonstrate here that targeting Stat3 or Smad7 for knockdown results in resensitization of TGF-β's cytostatic regulation in vivo. Overall, these results establish EGFR/Stat3/Smad7/TGF-β signaling axis driving tumor growth, which can be targeted therapeutically.
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Darcy A, Meltzer M, Miller J, Lee S, Chappell S, Ver Donck K, Montano M. A novel library screen identifies immunosuppressors that promote osteoblast differentiation. Bone 2012; 50:1294-303. [PMID: 22421346 PMCID: PMC3352976 DOI: 10.1016/j.bone.2012.03.001] [Citation(s) in RCA: 49] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 09/16/2011] [Revised: 02/29/2012] [Accepted: 03/02/2012] [Indexed: 12/22/2022]
Abstract
Bone homeostasis can be compromised by an increase in osteoclast-mediated resorption and/or a decrease in osteoblast-mediated bone deposition. While many efforts have focused on treating osteoclast resorption, there has been less emphasis on identifying strategies for promoting osteoblast function. Herein, we describe a high-throughput screening assay to select for small molecules that augment bone morphogenetic protein-2 (BMP-2)-mediated osteoblast lineage commitment. After an initial screen of 5405 compounds; consisting of FDA-approved drugs, known bioactives, and compounds with novel chemical makeup, we identified 45 small molecules that promoted osteoblast commitment. Of the 45 candidates, there was a broad array of classes that included nine retinoid analogs/derivatives and four immunosuppressants, notably rapamycin and FK-506, which were chosen for further study. Treatment of osteoblast precursor cells with rapamycin or FK-506, either alone, or synergistically with BMP-2, increased levels of phospho-Smad 1/5/8 protein and transcription of Runx-2, Osx and Smad-7, consistent with a role in promoting osteoblast differentiation. Only FK-506 was able to enhance osteocalcin transcripts and Alizarin Red staining, both late markers for differentiation. When osteoblast differentiation was suppressed with exogenous TGF-β1 treatment, rapamycin (but not FK-506) was able to rescue expression of differentiation markers, indicating distinct but overlapping activity of these compounds. Collectively, these data add to an understanding of pathways engaged in osteoblastogenesis, support a role for non-redundant immunosuppressant signaling, and provide a novel approach for the discovery of potentially therapeutic compounds that affect bone remodeling.
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Affiliation(s)
- Ariana Darcy
- Boston University School of Medicine, Boston, MA 02118, USA
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Mu Y, Gudey SK, Landström M. Non-Smad signaling pathways. Cell Tissue Res 2011; 347:11-20. [PMID: 21701805 DOI: 10.1007/s00441-011-1201-y] [Citation(s) in RCA: 432] [Impact Index Per Article: 30.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/17/2011] [Accepted: 06/03/2011] [Indexed: 12/19/2022]
Abstract
Transforming growth factor-beta (TGFβ) is a key regulator of cell fate during embryogenesis and has also emerged as a potent driver of the epithelial-mesenchymal transition during tumor progression. TGFβ signals are transduced by transmembrane type I and type II serine/threonine kinase receptors (TβRI and TβRII, respectively). The activated TβR complex phosphorylates Smad2 and Smad3, converting them into transcriptional regulators that complex with Smad4. TGFβ also uses non-Smad signaling pathways such as the p38 and Jun N-terminal kinase (JNK) mitogen-activated protein kinase (MAPK) pathways to convey its signals. Ubiquitin ligase tumor necrosis factor (TNF)-receptor-associated factor 6 (TRAF6) and TGFβ-associated kinase 1 (TAK1) have recently been shown to be crucial for the activation of the p38 and JNK MAPK pathways. Other TGFβ-induced non-Smad signaling pathways include the phosphoinositide 3-kinase-Akt-mTOR pathway, the small GTPases Rho, Rac, and Cdc42, and the Ras-Erk-MAPK pathway. Signals induced by TGFβ are tightly regulated and specified by post-translational modifications of the signaling components, since they dictate the subcellular localization, activity, and duration of the signal. In this review, we discuss recent findings in the field of TGFβ-induced responses by non-Smad signaling pathways.
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Affiliation(s)
- Yabing Mu
- Medical Biosciences, Umeå University, SE-901 85 Umeå, Sweden
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Abstract
TGF-β (transforming growth factor-β) is a pleiotropic cytokine regulating diverse cellular processes. It signals through membrane-bound receptors, downstream Smad proteins and/or other signalling mediators. Smad7 has been well established to be a key negative regulator of TGF-β signalling. It antagonizes TGF-β signalling through multiple mechanisms in the cytoplasm and in the nucleus. Smad7 can be transcriptionally induced by TGF-β and other growth factors and serves as an important cross-talk mediator of the TGF-β signalling pathway with other signalling pathways. Accordingly, it plays pivotal roles in embryonic development and adult homoeostasis, and altered expression of Smad7 is often associated with human diseases, such as cancer, tissue fibrosis and inflammatory diseases.
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Mordasky Markell L, Pérez-Lorenzo R, Masiuk KE, Kennett MJ, Glick AB. Use of a TGFbeta type I receptor inhibitor in mouse skin carcinogenesis reveals a dual role for TGFbeta signaling in tumor promotion and progression. Carcinogenesis 2010; 31:2127-35. [PMID: 20852150 DOI: 10.1093/carcin/bgq191] [Citation(s) in RCA: 26] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/16/2022] Open
Abstract
Pharmacological inhibitors of the transforming growth factor β (TGFβ) type I receptor (ALK5) have shown promise in blocking growth of xenotransplanted cancer cell lines but the effect on a multistage cancer model is not known. To test this, we treated mouse skin with SB431542 (SB), a well-characterized ALK5 inhibitor, during a two-stage skin carcinogenesis assay. Topical SB significantly reduced the total number, incidence and size of papillomas compared with 12-O-tetradecanoylphorbol 13-acetate (TPA) promotion alone, and this was linked to increased epidermal apoptosis, decreased proliferation and decreased cutaneous inflammation during promotion. In contrast, the frequency of conversion to squamous cell carcinoma (SCC) was 2-fold higher in papillomas treated with SB. Although there was no difference in tumor cell proliferation in early premalignant lesions, those that formed after SB treatment exhibited reduced squamous differentiation and an altered inflammatory microenvironment similar to SCC. In an inducible epidermal RAS transgenic model, treatment with SB enhanced proliferation and cutaneous inflammation in skin but decreased expression of keratin 1 and increased expression of simple epithelial keratin 18, markers of premalignant progression. In agreement with increased frequency of progression in the multistage model, SB treatment resulted in increased tumor formation with a more malignant phenotype following long-term RAS induction. In contrast to the current paradigm for TGFβ in carcinogenesis, these results demonstrate that cutaneous TGFβ signaling enables promotion of benign tumors but suppresses premalignant progression through context-dependent regulation of epidermal homeostasis and inflammation.
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Affiliation(s)
- Lauren Mordasky Markell
- Department of Veterinary and Biomedical Sciences, The Center for Molecular Toxicology and Carcinogenesis, Pennsylvania State University, University Park, PA 16802, USA
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Smad7 regulates the adult neural stem/progenitor cell pool in a transforming growth factor beta- and bone morphogenetic protein-independent manner. Mol Cell Biol 2010; 30:3685-94. [PMID: 20479122 DOI: 10.1128/mcb.00434-09] [Citation(s) in RCA: 23] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/12/2022] Open
Abstract
Members of the transforming growth factor beta (TGF-beta) family of proteins modulate the proliferation, differentiation, and survival of many different cell types. Neural stem and progenitor cells (NPCs) in the adult brain are inhibited in their proliferation by TGF-beta and by bone morphogenetic proteins (BMPs). Here, we investigated neurogenesis in a hypomorphic mouse model for the TGF-beta and BMP inhibitor Smad7, with the hypothesis that NPC proliferation might be reduced due to increased TGF-beta and BMP signaling. Unexpectedly, we found enhanced NPC proliferation as well as an increased number of label-retaining cells in vivo. The enhanced proliferation potential of mutant cells was retained in vitro in neurosphere cultures. We observed a higher sphere-forming capacity as well as faster growth and cell cycle progression. Use of specific inhibitors revealed that these effects were independent of TGF-beta and BMP signaling. The enhanced proliferation might be at least partially mediated by elevated signaling via epidermal growth factor (EGF) receptor, as mutant cells showed higher expression and activation levels of the EGF receptor. Conversely, an EGF receptor inhibitor reduced the proliferation of these cells. Our data indicate that endogenous Smad7 regulates neural stem/progenitor cell proliferation in a TGF-beta- and BMP-independent manner.
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Corpuz PS, Lindaman LL, Mellon PL, Coss D. FoxL2 Is required for activin induction of the mouse and human follicle-stimulating hormone beta-subunit genes. Mol Endocrinol 2010; 24:1037-51. [PMID: 20233786 PMCID: PMC2870942 DOI: 10.1210/me.2009-0425] [Citation(s) in RCA: 57] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/12/2009] [Accepted: 02/09/2010] [Indexed: 01/07/2023] Open
Abstract
Activin is a major physiological regulator of FSH. We identify FoxL2 as a critical component in activin induction of FSHbeta, both for the mouse gene, induction of which is Sma- and Mad-related protein (Smad) dependent, and for the human gene that is Smad independent. FoxL2 has been shown to regulate gonadotrope gene expression (GnRH receptor, alpha-glycoprotein subunit, porcine FSHbeta, and follistatin), but the mechanisms of action are not well understood. We identify novel sites required for activin action in both the mouse and human FSHbeta promoters, some of which bind FoxL2, and show that the FoxL2-binding element encompasses a larger region (12 bp) than the previously identified forkhead-binding consensus (7 bp). Remarkably, although required for activin induction, FoxL2 sites neither contribute to basal FSHbeta promoter activity nor confer activin response to a heterologous promoter; thus, they are neither classical activin-response elements nor is their role solely to recruit Smads to the promoter. FoxL2 overexpression can potentiate activin induction in gonadotropes and can confer activin responsiveness to FSHbeta in heterologous cells where this promoter is normally refractory to activin induction. Although Smad3 requires the presence of FoxL2 sites to induce mouse FSHbeta, even through its consensus Smad-binding element; the human promoter, which is induced by activin independently of Smad3, also requires FoxL2 sites for its induction by activin; thus the actions of FoxL2 are not exclusively through interactions with the Smad pathway. Thus, FoxL2 plays a key role in activin induction of the FSHbeta gene, by binding to sites conserved across multiple species.
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Affiliation(s)
- Patrick S Corpuz
- Department of Reproductive Medicine, University of California San Diego, 9500 Gilman Drive, La Jolla, California 92093-0674, USA
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Lu N, Carracedo S, Ranta J, Heuchel R, Soininen R, Gullberg D. The human α11 integrin promoter drives fibroblast-restricted expression in vivo and is regulated by TGF-β1 in a Smad- and Sp1-dependent manner. Matrix Biol 2010; 29:166-76. [DOI: 10.1016/j.matbio.2009.11.003] [Citation(s) in RCA: 29] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/10/2009] [Revised: 10/29/2009] [Accepted: 11/05/2009] [Indexed: 12/26/2022]
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Weiss A, Charbonnier E, Ellertsdóttir E, Tsirigos A, Wolf C, Schuh R, Pyrowolakis G, Affolter M. A conserved activation element in BMP signaling during Drosophila development. Nat Struct Mol Biol 2009; 17:69-76. [PMID: 20010841 DOI: 10.1038/nsmb.1715] [Citation(s) in RCA: 82] [Impact Index Per Article: 5.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/04/2009] [Accepted: 10/08/2009] [Indexed: 12/25/2022]
Abstract
The transforming growth factor beta (TGF-beta) family member Decapentaplegic (Dpp) is a key regulator of patterning and growth in Drosophila development. Previous studies have identified a short DNA motif called the silencer element (SE), which recruits a trimeric Smad complex and the repressor Schnurri to downregulate target enhancers upon Dpp signaling. We have now isolated the minimal enhancer of the dad gene and discovered a short motif we termed the activating element (AE). The AE is similar to the SE and recruits the Smad proteins via a conserved mechanism. However, the AE and SE differ at important nucleotide positions. As a consequence, the AE does not recruit Schnurri but rather integrates repressive input by the default repressor Brinker and activating input by the Smad signal transducers Mothers against Dpp (Mad) and Medea via competitive DNA binding. The AE allows the identification of hitherto unknown direct Dpp targets and is functionally conserved in vertebrates.
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Abstract
Transforming growth factor beta (TGFbeta) pathways are implicated in metazoan development, adult homeostasis and disease. TGFbeta ligands signal via receptor serine/threonine kinases that phosphorylate, and activate, intracellular Smad effectors as well as other signaling proteins. Oligomeric Smad complexes associate with chromatin and regulate transcription, defining the biological response of a cell to TGFbeta family members. Signaling is modulated by negative-feedback regulation via inhibitory Smads. We review here the mechanisms of TGFbeta signal transduction in metazoans and emphasize events crucial for embryonic development.
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Aoki MS, Soares AG, Miyabara EH, Baptista IL, Moriscot AS. Expression of genes related to myostatin signaling during rat skeletal muscle longitudinal growth. Muscle Nerve 2009; 40:992-9. [DOI: 10.1002/mus.21426] [Citation(s) in RCA: 31] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/08/2022]
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Abstract
Transforming growth factor (TGF)-β is a pleiotropic cytokine regulating a variety of cellular processes such as cell growth, differentiation, apoptosis, migration, cell adhesion, and immune response. In the well-understood classical TGF-β signaling pathway, TGF-β activates Smad signalling via its two cell surface receptors such as TβRII and ALK5/TβRI, leading to Smad-mediated transcriptional regulation. In addition, TGF-β may also activate other signaling pathways like mitogen-activated protein kinase, PI3K, etc. The signaling of TGF-β is finely regulated at different levels. Inhibitory Smads, including Smad6 and Smad7, are key regulators of TGF-β/bone morphogenetic protein (BMP) signaling by negative feedback loops. They can form stable complexes with activated type I receptors and thereby blocking the phosphorylation of R-Smads, or recruit ubiquitin E3 ligases, such as Smurf1/2, resulting in the ubiquitination and degradation of the activated type I receptors. Besides, these inhibitory Smad proteins also inhibit TGF-β/BMP signaling in the nucleus by interacting with transcriptional repressors, such as histone deacetylases, Hoxc-8, and CtBP, or disrupting the formation of the TGF-β-induced functional Smad-DNA complexes. Smad7 is in turn regulated by different stimuli, including TGF-β, IFN-γ, TNF-α as well as ultraviolet and TPA, and mediates the crosstalk between TGF-β and other signaling pathways. Deregulation of Smad7 expression has been associated with various human diseases, such as tissue fibrosis, inflammatory disease as well as carcinogenesis. Overexpression of Smad7 has been shown to antagonize TGF-β-mediated fibrosis, carcinogenesis, and inflammation, suggesting a therapeutic potential of Smad7 to treat these diseases.
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Affiliation(s)
- Xiaohua Yan
- State Key Laboratory of Biomembrane and Membrane Biotechnology, Department of Biological Sciences and Biotechnology, Tsinghua University, Beijing 100084, China
| | - Ziying Liu
- State Key Laboratory of Biomembrane and Membrane Biotechnology, Department of Biological Sciences and Biotechnology, Tsinghua University, Beijing 100084, China
| | - Yeguang Chen
- State Key Laboratory of Biomembrane and Membrane Biotechnology, Department of Biological Sciences and Biotechnology, Tsinghua University, Beijing 100084, China
- Correspondence address. Tel: +86-10-62795184; Fax: +86-10-62794376; E-mail:
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Abstract
Transforming growth factor-beta (TGF-beta)/bone morphogenic protein (BMP) signaling is involved in the vast majority of cellular processes and is fundamentally important during the entire life of all metazoans. Deregulation of TGF-beta/BMP activity almost invariably leads to developmental defects and/or diseases, including cancer. The proper functioning of the TGF-beta/BMP pathway depends on its constitutive and extensive communication with other signaling pathways, leading to synergistic or antagonistic effects and eventually desirable biological outcomes. The nature of such signaling cross-talk is overwhelmingly complex and highly context-dependent. Here we review the different modes of cross-talk between TGF-beta/BMP and the signaling pathways of Mitogen-activated protein kinase, phosphatidylinositol-3 kinase/Akt, Wnt, Hedgehog, Notch, and the interleukin/interferon-gamma/tumor necrosis factor-alpha cytokines, with an emphasis on the underlying molecular mechanisms.
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Affiliation(s)
- Xing Guo
- Department of Pharmacology and Cancer Biology, Duke University Medical Center, Durham, NC 27710, USA
| | - Xiao-Fan Wang
- Department of Pharmacology and Cancer Biology, Duke University Medical Center, Durham, NC 27710, USA
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Dancea HC, Shareef MM, Ahmed MM. Role of Radiation-induced TGF-beta Signaling in Cancer Therapy. MOLECULAR AND CELLULAR PHARMACOLOGY 2009; 1:44-56. [PMID: 20336170 PMCID: PMC2844640 DOI: 10.4255/mcpharmacol.09.06] [Citation(s) in RCA: 99] [Impact Index Per Article: 6.2] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Subscribe] [Scholar Register] [Indexed: 12/24/2022]
Abstract
TGF-β signaling regulates several different biological processes involving cell-growth, differentiation, apoptosis, motility, angiogenesis, epithelial mesenchymal transition and extracellular matrix production that affects embryonic development and pathogenesis of various diseases, including cancer, its effects depending on the cellular context and physiological environment. Growth suppression mediated by TGF-β signaling often associated with inhibition of c-myc, cdks and induction of p15, p27, Bax and p21. Despite its growth inhibitory effect, in certain conditions TGF-β may act as a promoter of cell proliferation and invasion. Loss of responsiveness to growth suppression by TGF-β due to mutation or loss of TGF-beta type II receptor (TβRII) and Smad4 in several different cancer cells are reported. In addition, TGF-β binding to its receptor activates many non-canonical signaling pathways. Radiation induced TGF-β is primarily involved in normal tissue injury and fibrosis. Seminal studies from our group have used radio-adjuvant therapies, involving classical components of the pathway such as TβRII and SMAD4 to overcome the growth promoting effects of TGF-β. The main impediment in the radiation-induced TGF-β signaling is the induction of SMAD7 that blocks TGF-β signaling in a negative feedback manner. It is well demonstrated from our studies that the use of neutralizing antibodies against TGF- β can render a robust radio-resistant effect. Thus, understanding the functional interactions of TGF-β signaling components of the pathway with other molecules may help tailor appropriate adjuvant radio-therapeutic strategies for treatment of solid tumors.
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Affiliation(s)
- Horatiu C Dancea
- Department of General Surgery, Geisinger Clinic, Danville, Pennsylvania
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Yu C, Azuma A, Li Y, Wang C, Abe S, Usuki J, Matsuda K, Kudoh S, Sunazuka T, Omura S. EM703, a new derivative of erythromycin, inhibits transforming growth factor-beta signaling in human lung fibroblasts. Exp Lung Res 2008; 34:343-54. [PMID: 18600500 DOI: 10.1080/01902140802093238] [Citation(s) in RCA: 11] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/21/2022]
Abstract
Long-term, low-dose macrolide therapy has been proven to improve survival in patients with diffuse panbronchiolitis and cystic fibrosis, although the mechanisms by which it does so remain unknown. To elucidate the molecular mechanisms of the anti-inflammatory effects of macrolides, the authors examined the effects of erythromycin (EM-A) and new derivative EM703 on transforming growth factor (TGF)-beta /Smad signaling fibroblasts. EM-A and EM703 each inhibited fibroblast proliferation and the collagen production in human lung fibroblasts induced by TGF-beta. EM-A and EM703 inhibited the augmentation of Smad3 mRNA induced by TGF-beta. Smad7 mRNA was inhibited by TGF-beta, but augmented by coincubation with EM-A or EM703. EM-A and EM703 each inhibited p-Smad2/3 proteins induced by TGF-beta. Smad7 protein inhibited by TGF-beta restored beyond basal level by EM-A and EM703. These findings suggest that EM-A and EM703 inhibit TGF-beta signaling in human lung fibroblasts via inhibition of p-Smad2/3 through recovery of Smad7 level.
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Affiliation(s)
- ChangHe Yu
- Department of Internal Medicine-Pulmonary Medicine, Infection, and Oncology, Nippon Medical School, Tokyo, Japan
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Moustakas A, Heldin CH. Dynamic control of TGF-β signaling and its links to the cytoskeleton. FEBS Lett 2008; 582:2051-65. [DOI: 10.1016/j.febslet.2008.03.027] [Citation(s) in RCA: 74] [Impact Index Per Article: 4.4] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/08/2008] [Accepted: 03/18/2008] [Indexed: 12/22/2022]
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Gohla G, Krieglstein K, Spittau B. Tieg3/Klf11 induces apoptosis in OLI-neu cells and enhances the TGF-β signaling pathway by transcriptional repression of Smad7. J Cell Biochem 2008; 104:850-61. [DOI: 10.1002/jcb.21669] [Citation(s) in RCA: 27] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/08/2022]
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Guo H, Leung JCK, Lam MF, Chan LYY, Tsang AWL, Lan HY, Lai KN. Smad7 transgene attenuates peritoneal fibrosis in uremic rats treated with peritoneal dialysis. J Am Soc Nephrol 2007; 18:2689-703. [PMID: 17855642 DOI: 10.1681/asn.2007010121] [Citation(s) in RCA: 40] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/18/2022] Open
Abstract
Transforming growth factor beta (TGF-beta) plays a critical role in the pathogenesis of the peritoneal fibrosis that complicates long-term peritoneal dialysis (PD). We studied the TGF-beta/Smad signaling pathway in peritoneal fibrosis induced in uremic rats treated with PD and explored the therapeutic potential of Smad7 to prevent fibrogenesis. After subtotal nephrectomy, uremic rats were treated with peritoneal dialysis using 4.25% dextrose-containing fluid. The peritoneum of uremic rats treated with PD demonstrated fibrosis, increased TGF-beta expression, increased Smad2/3 activation, decreased Smad7 expression, and increased expression of fibrogenic and angiogenic factors. In addition, peritoneal function was impaired and its structure was altered, including a thickened submesothelial layer. In rats transfected with a Smad7 transgene using an ultrasound-microbubble-mediated system, peritoneal fibrosis was attenuated, peritoneal function was improved, and Smad2/3 activation was inhibited. We suggest that administration of Smad7 inhibits peritoneal fibrogenesis in uremic rats treated with PD by correcting the imbalance between downregulated Smad7 and activated Smad2/3. Blockade of the TGF-beta/Smad signaling pathway may represent a novel therapeutic approach to prevent peritoneal fibrosis in patients treated with PD.
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Affiliation(s)
- Hong Guo
- Department of Medicine, Queen Mary Hospital, University of Hong Kong, Pokfulam, Hong Kong
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Pestov NB, Ahmad N, Korneenko TV, Zhao H, Radkov R, Schaer D, Roy S, Bibert S, Geering K, Modyanov NN. Evolution of Na,K-ATPase beta m-subunit into a coregulator of transcription in placental mammals. Proc Natl Acad Sci U S A 2007; 104:11215-20. [PMID: 17592128 PMCID: PMC2040879 DOI: 10.1073/pnas.0704809104] [Citation(s) in RCA: 24] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/18/2022] Open
Abstract
Change in gene functions (gene cooption) is one of the key mechanisms of molecular evolution. Genes can acquire new functions via alteration in properties of encoded proteins and/or via changes in temporal or spatial regulation of expression. Here we demonstrate radical changes in the functions of orthologous ATP1B4 genes during evolution of vertebrates. Expression of ATP1B4 genes is brain-specific in teleost fishes, whereas it is predominantly muscle-specific in tetrapods. The encoded beta m-proteins in fish, amphibian, and avian species are beta-subunits of Na,K-ATPase located in the plasma membrane. In placental mammals beta m-proteins lost their ancestral functions, accumulate in nuclear membrane of perinatal myocytes, and associate with transcriptional coregulator Ski-interacting protein (SKIP). Through interaction with SKIP, eutherian beta m acquired new functions as exemplified by regulation of TGF-beta-responsive reporters and by augmentation of mRNA levels of Smad7, an inhibitor of TGF-beta signaling. Thus, orthologous vertebrate ATP1B4 genes represent an instance of gene cooption that created fundamental changes in the functional properties of the encoded proteins.
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Affiliation(s)
- Nikolay B. Pestov
- *Department of Physiology and Pharmacology, University of Toledo College of Medicine, 3000 Arlington Avenue, Toledo, OH 43614
- Shemyakin–Ovchinnikov Institute of Bioorganic Chemistry, 16/10 Miklukho-Maklaya, Moscow 117997, Russia; and
| | - Nisar Ahmad
- *Department of Physiology and Pharmacology, University of Toledo College of Medicine, 3000 Arlington Avenue, Toledo, OH 43614
| | - Tatiana V. Korneenko
- *Department of Physiology and Pharmacology, University of Toledo College of Medicine, 3000 Arlington Avenue, Toledo, OH 43614
- Shemyakin–Ovchinnikov Institute of Bioorganic Chemistry, 16/10 Miklukho-Maklaya, Moscow 117997, Russia; and
| | - Hao Zhao
- *Department of Physiology and Pharmacology, University of Toledo College of Medicine, 3000 Arlington Avenue, Toledo, OH 43614
| | - Rossen Radkov
- *Department of Physiology and Pharmacology, University of Toledo College of Medicine, 3000 Arlington Avenue, Toledo, OH 43614
| | - Danièle Schaer
- Department of Pharmacology and Toxicology, University of Lausanne, Rue du Bugnon 27, CH-1005 Lausanne, Switzerland
| | - Sophie Roy
- Department of Pharmacology and Toxicology, University of Lausanne, Rue du Bugnon 27, CH-1005 Lausanne, Switzerland
| | - Stéphanie Bibert
- Department of Pharmacology and Toxicology, University of Lausanne, Rue du Bugnon 27, CH-1005 Lausanne, Switzerland
| | - Käthi Geering
- Department of Pharmacology and Toxicology, University of Lausanne, Rue du Bugnon 27, CH-1005 Lausanne, Switzerland
| | - Nikolai N. Modyanov
- *Department of Physiology and Pharmacology, University of Toledo College of Medicine, 3000 Arlington Avenue, Toledo, OH 43614
- To whom correspondence should be addressed at:
Department of Physiology and Pharmacology, University of Toledo College of Medicine, Health Science Campus, 3000 Arlington Avenue, Mail Stop 1008, Toledo, OH 43614-2598. E-mail:
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