A Kinase Domain-truncated Type I Receptor Blocks Bone Morphogenetic Protein-2-induced Signal Transduction in C2C12 Myoblasts

Namiki, Mana; Akiyama, Shinichiro; Katagiri, Takenobu; Suzuki, Atsushi; Ueno, Naoto; Yamaji, Noboru; Rosen, Vicki; Wozney, John M.; Suda, Toshio

doi:10.1074/jbc.272.35.22046

Cited by 78 publications

(56 citation statements)

References 53 publications

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“…TGF-␤ and BMP-2 block the myogenic differentiation of C2C12 cells by suppressing the expression of master control genes, including that of MyoD (20,35). Our data show that overexpression of Runx2 suppresses MyoD expression and stops the myogenic differentiation of C2C12 cells (Fig.…”

Section: Discussionmentioning

confidence: 54%

Runx2 Is a Common Target of Transforming Growth Factor β1 and Bone Morphogenetic Protein 2, and Cooperation between Runx2 and Smad5 Induces Osteoblast-Specific Gene Expression in the Pluripotent Mesenchymal Precursor Cell Line C2C12

Lee

Kim

et al. 2000

Molecular and Cellular Biology

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Transforming growth factor ␤ (TGF-␤) is a potent multifunctional regulator of cell growth and differentiation. Although nearly all cells synthesize and respond to TGF-␤, bone and cartilage are particularly rich in this growth factor (6, 46). TGF-␤1, the prototypic member of the TGF-␤ superfamily, elicits diverse cellular responses, including (i) inhibition of adipogenesis and myogenesis and (ii) stimulation of chondrogenesis and osteogenesis (31). TGF-␤1 stimulates the synthesis of matrix proteins and their receptors (for example, fibronectin, fibronectin receptor, collagen, osteonectin, osteopontin, and integrins) and inhibits matrix degradation by increasing the production of protease inhibitors and decreasing the production of proteases (42). Members of the TGF-␤ superfamily with important effects on bone cell differentiation are bone morphogenetic proteins (BMPs) (17, 41), which were first identified as factors that induce bone formation in vivo when implanted into muscular tissues (54). Unlike TGF-␤, which induces new bone formation only when injected near bone, BMPs produce bone formation even when injected into ectopic sites. TGF-␤ and BMPs bind to distinct receptors, TGF-␤ type I and II receptors for TGF-␤ and BMP type I and II receptors for BMPs. Following ligand binding, the receptor-associated kinase is activated and phosphorylates Smads, which move into the nucleus to stimulate the transcription of a set of target genes. Smad2 and -3 are activated by TGF-␤ receptors and mediate TGF-␤ responses, whereas Smad1, -5, and -8 are activated by BMP receptors and transduce BMP signals (15,32,57).The pluripotent mesenchymal precursor cell line C2C12 provides a model system to study the early stage of osteoblast differentiation during bone formation in muscular tissues. In this model, TGF-␤1 inhibits the differentiation of C2C12 cells into multinucleated myotubes without inducing osteoblast phenotypes. BMP-2 not only inhibits the terminal differentiation of C2C12 cells but also induces osteoblast phenotypes (20). Therefore, the C2C12 model is useful for analyzing both the common and specific signaling mechanisms of TGF-␤ and BMPs. In C2C12 cells, overexpression of Smad1 and Smad5 induced alkaline phosphatase (ALP) activity, a typical osteoblast-specific marker, and inhibited muscle-specific gene expression (11,36,56). These results suggested that BMP functions via either Smad1 or Smad5 and that the induction of the osteoblast phenotype and the inhibition of myogenic differentiation are regulated at the transcriptional level. However, the molecular mechanisms through which Smads block myogenic differentiation and induce osteogenic differentiation are not known.Runx/PEBP2/Cbf (hereafter referred to as Runx) is a sequence-specific DNA binding protein that recognizes a specific DNA sequence originally identified as the binding site for

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Section: Discussionmentioning

confidence: 54%

Runx2 Is a Common Target of Transforming Growth Factor β1 and Bone Morphogenetic Protein 2, and Cooperation between Runx2 and Smad5 Induces Osteoblast-Specific Gene Expression in the Pluripotent Mesenchymal Precursor Cell Line C2C12

Lee

Kim

et al. 2000

Molecular and Cellular Biology

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“…Alkaline Phosphatase Staining-Alkaline phosphatase (ALP) activity has been widely accepted as a simple and easiest way to determine the osteogenic differentiation, especially in the BMP-2-induced osteogenic transdifferentiation in C2C12 cells (3,6,7). Cells were washed with phosphate-buffered saline twice, fixed with 2% paraformaldehyde, and stained for alkaline phosphatase according to the manufacturer's instructions (Sigma).…”

Section: Methodsmentioning

confidence: 99%

“…BMPR-I is further subclassified into BMPR-IA (also called ALK3) and BMPR-IB (also called ALK6). It has been shown that both the inhibition of myoblast differentiation and the induction of osteoblast differentiation by BMP-2 involve the activations of BMPR-I receptors (6,7), their intracellular transducers Smad1 and Smad5 (8,9), and the osteogenic master transcription factor Runx2 (4,10).…”

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confidence: 99%

BMP-2-induced Runx2 Expression Is Mediated by Dlx5, and TGF-β1 Opposes the BMP-2-induced Osteoblast Differentiation by Suppression of Dlx5 Expression

Lee

Kim

et al. 2003

Journal of Biological Chemistry

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“…cells, BMP-6 mainly utilizes ALK-2, whereas BMP-2 utilizes ALK-3 for signaling (19), because ALK-6 is not expressed in these cells (19,30). We thus examined the effects of I-Smads on BMP-6-and BMP-2-induced signaling in C2C12 cells (Fig.…”

Section: Effect Of Smad6 On Bmp-6-induced Alk-2 Activation and Bmp-2-mentioning

confidence: 99%

Selective Inhibitory Effects of Smad6 on Bone Morphogenetic Protein Type I Receptors

Goto

Kamiya

Imamura

et al. 2007

Journal of Biological Chemistry

128

108

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The inhibitory Smads, Smad6 and Smad7, play pivotal roles in negative regulation of transforming growth factor-␤ (TGF-␤) 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-␤/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.

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A Kinase Domain-truncated Type I Receptor Blocks Bone Morphogenetic Protein-2-induced Signal Transduction in C2C12 Myoblasts

Abstract: Members of the transforming growth factor (TGF)-

Cited by 78 publications

References 53 publications

Runx2 Is a Common Target of Transforming Growth Factor β1 and Bone Morphogenetic Protein 2, and Cooperation between Runx2 and Smad5 Induces Osteoblast-Specific Gene Expression in the Pluripotent Mesenchymal Precursor Cell Line C2C12

Runx2 Is a Common Target of Transforming Growth Factor β1 and Bone Morphogenetic Protein 2, and Cooperation between Runx2 and Smad5 Induces Osteoblast-Specific Gene Expression in the Pluripotent Mesenchymal Precursor Cell Line C2C12

BMP-2-induced Runx2 Expression Is Mediated by Dlx5, and TGF-β1 Opposes the BMP-2-induced Osteoblast Differentiation by Suppression of Dlx5 Expression

Selective Inhibitory Effects of Smad6 on Bone Morphogenetic Protein Type I Receptors

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