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Roles of bone morphogenetic protein type I receptors and Smad proteins in osteoblast and chondroblast differentiation 总被引:18,自引:0,他引:18
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Fujii M Takeda K Imamura T Aoki H Sampath TK Enomoto S Kawabata M Kato M Ichijo H Miyazono K 《Molecular biology of the cell》1999,10(11):3801-3813
The biological effects of type I serine/threonine kinase receptors and Smad proteins were examined using an adenovirus-based vector system. Constitutively active forms of bone morphogenetic protein (BMP) type I receptors (BMPR-IA and BMPR-IB; BMPR-I group) and those of activin receptor-like kinase (ALK)-1 and ALK-2 (ALK-1 group) induced alkaline phosphatase activity in C2C12 cells. Receptor-regulated Smads (R-Smads) that act in the BMP pathways, such as Smad1 and Smad5, also induced the alkaline phosphatase activity in C2C12 cells. BMP-6 dramatically enhanced alkaline phosphatase activity induced by Smad1 or Smad5, probably because of the nuclear translocation of R-Smads triggered by the ligand. Inhibitory Smads, i.e., Smad6 and Smad7, repressed the alkaline phosphatase activity induced by BMP-6 or the type I receptors. Chondrogenic differentiation of ATDC5 cells was induced by the receptors of the BMPR-I group but not by those of the ALK-1 group. However, kinase-inactive forms of the receptors of the ALK-1 and BMPR-I groups blocked chondrogenic differentiation. Although R-Smads failed to induce cartilage nodule formation, inhibitory Smads blocked it. Osteoblast differentiation induced by BMPs is thus mediated mainly via the Smad-signaling pathway, whereas chondrogenic differentiation may be transmitted by Smad-dependent and independent pathways. 相似文献
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Valcourt U Gouttenoire J Moustakas A Herbage D Mallein-Gerin F 《The Journal of biological chemistry》2002,277(37):33545-33558
We investigated the effects of bone morphogenetic protein (BMP)-2, a member of the transforming growth factor-beta superfamily, on the regulation of the chondrocyte phenotype, and we identified signaling molecules involved in this regulation. BMP-2 triggers three concomitant responses in mouse primary chondrocytes and chondrocytic MC615 cells. First, BMP-2 stimulates expression or synthesis of type II collagen. Second, BMP-2 induces expression of molecular markers characteristic of pre- and hypertrophic chondrocytes, such as Indian hedgehog, parathyroid hormone/parathyroid hormone-related peptide receptor, type X collagen, and alkaline phosphatase. Third, BMP-2 induces osteocalcin expression, a specific trait of osteoblasts. Constitutively active forms of transforming growth factor-beta family type I receptors and Smad proteins were overexpressed to address their role in this process. Activin receptor-like kinase (ALK)-1, ALK-2, ALK-3, and ALK-6 were able to reproduce the hypertrophic maturation of chondrocytes induced by BMP-2. In addition, ALK-2 mimicked further the osteoblastic differentiation of chondrocytes induced by BMP-2. In the presence of BMP-2, Smad1, Smad5, and Smad8 potentiated the hypertrophic maturation of chondrocytes, but failed to induce osteocalcin expression. Smad6 and Smad7 impaired chondrocytic expression and osteoblastic differentiation induced by BMP-2. Thus, our results indicate that Smad-mediated pathways are essential for the regulation of the different steps of chondrocyte and osteoblast differentiation and suggest that additional Smad-independent pathways might be activated by ALK-2. 相似文献
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Selective inhibitory effects of Smad6 on bone morphogenetic protein type I receptors 总被引:1,自引:0,他引:1
Goto K Kamiya Y Imamura T Miyazono K Miyazawa K 《The Journal of biological chemistry》2007,282(28):20603-20611
The inhibitory Smads, Smad6 and Smad7, play pivotal roles in negative regulation of transforming growth factor-beta (TGF-beta) family signaling as feedback molecules as well as mediators of cross-talk with other signaling pathways. Whereas Smad7 acts as a ubiquitous inhibitor of Smad signaling, Smad6 has been shown to effectively inhibit bone morphogenetic protein (BMP) signaling but only weakly TGF-beta/activin signaling. In the present study, we have found that Smad6 inhibits signaling from the activin receptor-like kinase (ALK)-3/6 subgroup in preference to that from the ALK-1/2 subgroup of BMP type I receptors. The difference is attributable to the interaction of Smad6 with these BMP type I receptors. The amino acid residues responsible for Smad6 sensitivity of ALK-3 were identified as Arg-238, Phe-264, Thr-265, and Ala-269, which map to the N-terminal lobe of the ALK-3 kinase domain. Although Smad6 regulates BMP signaling through multiple mechanisms, our findings suggest that interaction with type I receptors is a critical step in the function of Smad6. 相似文献
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In mammals, two inhibitory Smads (I-Smads), Smad6 and Smad7, play pivotal roles in negative regulation of TGF-beta family signaling. Smad7 ubiquitously inhibits TGF-beta family signaling, whereas Smad6 inhibits signaling from the ALK-3/6 subfamily in preference to that from the ALK-1/2 and ALK-4/5/7 subfamilies of TGF-beta family type I receptors. In Drosophila, only one I-Smad, Dad, has been identified. Here we examined inhibitory effects of Dad on type I receptors in Drosophila. Dad inhibited Saxophone (ALK-1/2 orthologue) and Thickveins (ALK-3/6 orthologue) but not Baboon (ALK-4/5/7 orthologue). The differential modes of action of I-Smads in mammals and Drosophila are discussed. 相似文献
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Activin, a member of the transforming growth factor beta (TGF-beta) superfamily, signals through a heteromeric complex of type I and type II serine-threonine kinase receptors. The two activin type I receptors previously identified, ALK-2 (ActR-I) and ALK-4 (ActR-IB), have distinct effects on gene expression, differentiation and morphogenesis in the Xenopus animal cap assay. ALK-4 reproduces the effects of activin treatment including the dose-dependent induction of progressively more dorso-anterior mesodermal and endodermal markers, whereas ALK-2 induces only ventral mesodermal markers and counteracts the effects of ALK-4. To identify regions of the receptors that determine signaling specificity we have generated chimeras of the constitutively active ALK-2 and ALK-4 receptors (termed ALK-2* and ALK-4*). The effects of these chimeric receptors on gene expression and morphogenetic movements implicate the loop between kinase subdomains IV and V in mediating the strong dorsal gene-inducing properties of ALK-4*; when the seven amino acids comprising this loop are transferred from ALK-4* to ALK-2*, the resulting chimeric receptor is capable of inducing the expression of dorsal-specific genes. In contrast, when the equivalent region of ALK-2* is transferred to the ALK-4* backbone it cannot effectively counteract the dorsalizing effects of ALK-4*, suggesting that other regions of type I receptors are also involved in determining signal specificity. 相似文献
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Functional roles of the bone morphogenetic protein system in thyrotropin signaling in porcine thyroid cells 总被引:2,自引:0,他引:2
Suzuki J Otsuka F Takeda M Inagaki K Miyoshi T Mimura Y Ogura T Doihara H Makino H 《Biochemical and biophysical research communications》2005,327(4):1124-1130
We uncovered a new regulation of thyrocyte function by bone morphogenetic protein (BMP) under the influence of thyrotropin (TSH) using primary culture of porcine thyrocytes. The BMP type I receptors, ALK-2 (ActRIA), -3 (BMPRIA), and -6 (BMPRIB), were expressed in porcine thyrocytes, while ALK-6 was not detected in human thyroid. Treatment with BMP-2, -4, -6, -7, and TGF-beta1 exhibited a dose-dependent suppression of DNA synthesis by porcine thyrocytes. BMP-2, -4, -6, -7, and TGF-beta1 suppressed TSH receptor mRNA expression on thyrocytes, which was consistent with their suppressive effect on TSH-induced cAMP synthesis and TSH-induced insulin-like growth factor-1 expression. Activin exhibited minimal suppression of thyrocyte DNA synthesis and did not exhibit suppressive effects on TSH receptor mRNA expression. Phosphorylated Smad1/5/8 was detected in the lysates of porcine thyrocytes treated with BMP-2, -4, -6, and -7. However, in the presence of TSH, BMP-6 and -7 failed to activate Smad1/5/8 phosphorylation and 3TP-reporter activity, whereas BMP-2 and -4 maintained clear activation of the BMP signaling regardless of the presence of TSH. This diverged regulation of thyroid BMP system by TSH is most likely due to the reduction of ALK-6 expression caused by TSH. Thus, the thyroid BMP system is functionally linked to TSH actions through modulating TSH receptor expression and TSH, in turn, selectively inhibits BMP signaling. Given that BMP system is present in human thyroid and the expression pattern of ALK-2 and BMPRII is different between follicular adenomas and normal thyroid tissues, the endogenous BMP system may be involved in regulating thyrocyte growth and TSH sensitivity of human thyroid adenomas. 相似文献
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Xiying Wu Claudius E. Robinson Hon Wa Fong Jeffrey M. Gimble 《Journal of cellular physiology》1996,168(2):453-461
The bone morphogenetic proteins, members of the transforming growth factor-β cytokine family, induce the osteoblast phenotype and promote osteogenesis in the bone marrow stroma. Simultaneously, these cytokines inhibit other mesodermal differentiation pathways, such as adipogenesis and myogenesis. The receptors for the bone morphogenetic proteins belong to a family of transmembrane serine/threonine kinase TGFβ type I and type II receptor proteins. In man, these include the activin receptor like kinase-3 (ALK-3), a type I receptor protein. We have used a polyclonal antibody to examine the expression of the native murine ALK-3 protein in murine tissues and bone morphogenetic protein-responsive cell lines. On Western blot analyses, we found that the native 85 kDa native ALK-3 protein was expressed in a number of murine tissues; protein and mRNA levels did not necessarily correlate. Two bone morphogenetic protein-responsive cell lines, BMS2 bone marrow stromal cells and C2C12 myoblasts, expressed the ALK-3 protein constitutively. Cell differentiation was accompanied by modest changes in ALK-3 protein levels. Immunoprecipitation of the ALK-3 protein cross linked to [125I] BMP-4 revealed two major receptor complexes of approximately 90 kDa and 170 kDa in size. Biotin surface-labeling experiments revealed that the 85 kDa ALK-3 protein was constitutively associated with a novel 140 kDa surface glycoprotein. Deglycosylation reduced the protein's size to 116 kDa, comparable in size to that of the recently described BMP type II receptor. These findings support the current model that BMP interacts with a pre-existing complex consisting of a type I and type II receptor protein. © 1996 Wiley-Liss, Inc. 相似文献
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Nishihara A Fujii M Sampath TK Miyazono K Reddi AH 《Biochemical and biophysical research communications》2003,301(2):617-622
Articular chondrocytes progressively undergo dedifferentiation into a spindle-shaped mesenchymal cellular phenotype in monolayers. Chondrocyte dedifferentiation is stimulated by retinoic acid. On the other hand, bone morphogenic proteins (BMPs) stimulate differentiation of chondrocytes. We examined the mechanism of effects of BMP in chondrocyte differentiation with use of a recombinant adenovirus vector system. Constitutively active forms of BMP type I receptors (BMPR-IA and BMPR-IB) and those of activin receptor-like kinase (ALK)-1 and ALK-2 maintained differentiation of chondrocytes in the presence of retinoic acid. The BMP receptor-regulated signaling substrates, Smad1/5, weakly induced chondrocyte differentiation; the effects of Smad1/5 were enhanced by BMP-7 treatment. Inhibitory Smad, Smad6, blocked increase of expression of chondrocyte markers by BMP-7 in a dose-dependent manner. SB202190, a p38 mitogen-activated protein kinase inhibitor, inhibited this effect of BMP-7; however, since SB202190 suppressed phosphorylation of Smad1/5, this may be due to blockade of BMP receptor activation. These results together strongly suggest that induction of chondrocyte differentiation by BMP-7 is regulated by Smad pathways. 相似文献
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Fukuda T Kohda M Kanomata K Nojima J Nakamura A Kamizono J Noguchi Y Iwakiri K Kondo T Kurose J Endo K Awakura T Fukushi J Nakashima Y Chiyonobu T Kawara A Nishida Y Wada I Akita M Komori T Nakayama K Nanba A Maruki Y Yoda T Tomoda H Yu PB Shore EM Kaplan FS Miyazono K Matsuoka M Ikebuchi K Ohtake A Oda H Jimi E Owan I Okazaki Y Katagiri T 《The Journal of biological chemistry》2009,284(11):7149-7156
Fibrodysplasia ossificans progressiva (FOP) is a rare autosomal dominant disorder characterized by congenital malformation of the great toes and by progressive heterotopic bone formation in muscle tissue. Recently, a mutation involving a single amino acid substitution in a bone morphogenetic protein (BMP) type I receptor, ALK2, was identified in patients with FOP. We report here that the identical mutation, R206H, was observed in 19 Japanese patients with sporadic FOP. This mutant receptor, ALK2(R206H), activates BMP signaling without ligand binding. Moreover, expression of Smad1 and Smad5 was up-regulated in response to muscular injury. ALK2(R206H) with Smad1 or Smad5 induced osteoblastic differentiation that could be inhibited by Smad7 or dorsomorphin. Taken together, these findings suggest that the heterotopic bone formation in FOP may be induced by a constitutively activated BMP receptor signaling through Smad1 or Smad5. Gene transfer of Smad7 or inhibition of type I receptors with dorsomorphin may represent strategies for blocking the activity induced by ALK2(R206H) in FOP. 相似文献
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Scherner O Meurer SK Tihaa L Gressner AM Weiskirchen R 《The Journal of biological chemistry》2007,282(19):13934-13943
Transforming growth factor-beta1 (TGF-beta1) and BMP-7 (bone morphogenetic protein-7; OP-1) play central, antagonistic roles in kidney fibrosis, a setting in which the expression of endoglin (CD105), an accessory TGF-beta type III receptor, is increased. So far, endoglin is known as a negative regulator of TGF-beta/ALK-5 signaling. Here we analyzed the effect of BMP-7 on TGF-beta1 signaling and the role of endoglin for both pathways in endoglin-deficient L(6)E(9) cells. In this myoblastic cell line, TGF-beta1 and BMPs are opposing cytokines, interfering with myogenic differentiation. Both induce specific target genes of which Id1 (for BMPs) and collagen I (for TGF-beta1) are two examples. TGF-beta1 activated two distinct type I receptors, ALK-5 and ALK-1, in these cells. Although the ALK-5/Smad3 signaling pathway mediated collagen I expression, ALK-1/Smad1/Smad5 signaling mediated a transient Id1 up-regulation. In contrast, BMP-7 exclusively activated Smad1/Smad5 resulting in a more prolonged Id1 expression. Although BMP-7 had no impact on collagen I abundance, it antagonized TGF-beta1-induced collagen I expression and (CAGA)(12)-MLP-Luc activity, effects that are mediated by the ALK-5/Smad3 pathway. Finally, we found that the transient overexpression of endoglin, previously shown to inhibit TGF-beta1-induced ALK-5/Smad3 signaling, enhanced the BMP-7/Smad1/Smad5 pathway. 相似文献
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Mazerbourg S Sangkuhl K Luo CW Sudo S Klein C Hsueh AJ 《The Journal of biological chemistry》2005,280(37):32122-32132
There are more than 30 human transforming growth factor beta/bone morphogenetic protein/growth differentiation factor (TGFbeta/BMP/GDF)-related ligands known to be important during embryonic development, organogenesis, bone formation, reproduction, and other physiological processes. Although select TGFbeta/BMP/GDF proteins were found to interact with type II and type I serine/threonine receptors to activate downstream Smad and other proteins, the receptors and signaling pathways for one-third of these TGFbeta/BMP/GDF paralogs are still unclear. Based on a genomic analysis of the entire repertoire of TGFbeta/BMP/GDF ligands and serine/threonine kinase receptors, we tested the ability of three orphan BMP/GDF ligands to activate a limited number of phylogenetically related receptors. We characterized the dimeric nature of recombinant GDF6 (also known as BMP13), GDF7 (also known as BMP12), and BMP10. We demonstrated their bioactivities based on the activation of Smad1/5/8-, but not Smad2/3-, responsive promoter constructs in the MC3T3 cell line. Furthermore, we showed their ability to induce the phosphorylation of Smad1, but not Smad2, in these cells. In COS7 cells transfected with the seven known type I receptors, overexpression of ALK3 or ALK6 conferred ligand signaling by GDF6, GDF7, and BMP10. In contrast, transfection of MC3T3 cells with ALK3 small hairpin RNA suppressed Smad signaling induced by all three ligands. Based on the coevolution of ligands and receptors, we also tested the role of BMPRII and ActRIIA as the type II receptor candidates for the three orphan ligands. We found that transfection of small hairpin RNA for BMPRII and ActRIIA in MC3T3 cells suppressed the signaling of GDF6, GDF7, and BMP10. Thus, the present approach provides a genomic paradigm for matching paralogous polypeptide ligands with a limited number of evolutionarily related receptors capable of activating specific downstream Smad proteins. 相似文献
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Cooperative inhibition of bone morphogenetic protein signaling by Smurf1 and inhibitory Smads 总被引:9,自引:0,他引:9
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Murakami G Watabe T Takaoka K Miyazono K Imamura T 《Molecular biology of the cell》2003,14(7):2809-2817
Smad ubiquitin regulatory factor (Smurf) 1 binds to receptor-regulated Smads for bone morphogenetic proteins (BMPs) Smad1/5 and promotes their degradation. In addition, Smurf1 associates with transforming growth factor-beta type I receptor through the inhibitory Smad (I-Smad) Smad7 and induces their degradation. Herein, we examined whether Smurf1 negatively regulates BMP signaling together with the I-Smads Smad6/7. Smurf1 and Smad6 cooperatively induced secondary axes in Xenopus embryos. Using a BMP-responsive promoter-reporter construct in mammalian cells, we found that Smurf1 cooperated with I-Smad in inhibiting BMP signaling and that the inhibitory activity of Smurf1 was not necessarily correlated with its ability to bind to Smad1/5 directly. Smurf1 bound to BMP type I receptors via I-Smads and induced ubiquitination and degradation of these receptors. Moreover, Smurf1 associated with Smad1/5 indirectly through I-Smads and induced their ubiquitination and degradation. Smurf1 thus controls BMP signaling with and without I-Smads through multiple mechanisms. 相似文献
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Alejandre-Alcázar MA Kwapiszewska G Reiss I Amarie OV Marsh LM Sevilla-Pérez J Wygrecka M Eul B Köbrich S Hesse M Schermuly RT Seeger W Eickelberg O Morty RE 《American journal of physiology. Lung cellular and molecular physiology》2007,292(2):L537-L549
Prematurely born infants who require oxygen therapy often develop bronchopulmonary dysplasia (BPD), a debilitating disorder characterized by pronounced alveolar hypoplasia. Hyperoxic injury is believed to disrupt critical signaling pathways that direct lung development, causing BPD. We investigated the effects of normobaric hyperoxia on transforming growth factor (TGF)-beta and bone morphogenetic protein (BMP) signaling in neonatal C57BL/6J mice exposed to 21% or 85% O(2) between postnatal days P1 and P28. Growth and respiratory compliance were significantly impaired in pups exposed to 85% O(2), and these pups also exhibited a pronounced arrest of alveolarization, accompanied by dysregulated expression and localization of both receptor (ALK-1, ALK-3, ALK-6, and the TGF-beta type II receptor) and Smad (Smads 1, 3, and 4) proteins. TGF-beta signaling was potentiated, whereas BMP signaling was impaired both in the lungs of pups exposed to 85% O(2) as well as in MLE-12 mouse lung epithelial cells and NIH/3T3 and primary lung fibroblasts cultured in 85% O(2). After exposure to 85% O(2), primary alveolar type II cells were more susceptible to TGF-beta-induced apoptosis, whereas primary pulmonary artery smooth muscle cells were unaffected. Exposure of primary lung fibroblasts to 85% O(2) significantly enhanced the TGF-beta-stimulated production of the alpha(1) subunit of type I collagen (Ialpha(1)), tissue inhibitor of metalloproteinase-1, tropoelastin, and tenascin-C. These data demonstrated that hyperoxia significantly affects TGF-beta/BMP signaling in the lung, including processes central to septation and, hence, alveolarization. The amenability of these pathways to genetic and pharmacological manipulation may provide alternative avenues for the management of BPD. 相似文献
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Nonaka K Shum L Takahashi I Takahashi K Ikura T Dashner R Nuckolls GH Slavkin HC 《The International journal of developmental biology》1999,43(8):795-807
Bone morphogenetic protein 4 (BMP4) induces, whereas epidermal growth factor (EGF) inhibits chondrogenesis. We hypothesize that BMP4 and EGF mediated intracellular signals are both coupled in the regulation of Meckel's cartilage development. Two chondrogenic experimental model systems were employed to test the hypothesis: (1) an ex vivo, serum-free, organ culture system for mouse embryonic mandibular processes, and (2) a micromass culture system for chicken embryonic mandibular processes. Chondrogenesis was assayed by alcian blue staining and expression of Sox9 and type II collagen. Exogenous EGF inhibited and BMP4 induced ectopic cartilage in a dose-dependent manner. When BMP4- and EGF-soaked beads were implanted in juxtaposition within embryonic day 10 mouse mandibular processes, the incidence and amount of ectopic cartilage, and Sox9 and type II collagen expression induced by BMP4, were significantly reduced as the concentration of EGF was increased. Similarly, in chicken serum-free micromass cultures, expression of a constitutively active BMP receptor type IB by replication competent avian retrovirus system promoted the rate and extent of chondrogenesis; however, exogenous EGF attenuated this effect. In micromass cultures, BMP signaling resulted in nuclear translocation and accumulation of the signaling molecule Smad1, whereas the addition of EGF inhibited this event. Our results suggest that BMP4 and EGF function antagonistically, yet are coupled in the regulation of initial chondrogenesis. Smad1 serves as a point of convergence for the integration of two different growth factor signaling pathways during chondrogenesis. 相似文献