RESUMO
Myostatin, a transforming growth factor beta (TGF-beta) family member, is a potent negative regulator of skeletal muscle growth. In this study we characterized the myostatin signal transduction pathway and examined its effect on bone morphogenetic protein (BMP)-induced adipogenesis. While both BMP7 and BMP2 activated transcription from the BMP-responsive I-BRE-Lux reporter and induced adipogenic differentiation, myostatin inhibited BMP7- but not BMP2-mediated responses. To dissect the molecular mechanism of this antagonism, we characterized the myostatin signal transduction pathway. We showed that myostatin binds the type II Ser/Thr kinase receptor. ActRIIB, and then partners with a type I receptor, either activin receptor-like kinase 4 (ALK4 or ActRIB) or ALK5 (TbetaRI), to induce phosphorylation of Smad2/Smad3 and activate a TGF-beta-like signaling pathway. We demonstrated that myostatin prevents BMP7 but not BMP2 binding to its receptors and that BMP7-induced heteromeric receptor complex formation is blocked by competition for the common type II receptor, ActRIIB. Thus, our results reveal a strikingly specific antagonism of BMP7-mediated processes by myostatin and suggest that myostatin is an important regulator of adipogenesis.
Assuntos
Adipócitos/metabolismo , Transdução de Sinais , Fator de Crescimento Transformador beta/metabolismo , Animais , Proteína Morfogenética Óssea 2 , Proteína Morfogenética Óssea 7 , Proteínas Morfogenéticas Ósseas/metabolismo , Células COS , Diferenciação Celular , Linhagem Celular , Proteínas de Ligação a DNA/metabolismo , Genes Reporter , Homeostase , Humanos , Camundongos , Camundongos Endogâmicos C3H , Modelos Biológicos , Miostatina , Células NIH 3T3 , Fosforilação , Reação em Cadeia da Polimerase , Ligação Proteica , RNA/metabolismo , Interferência de RNA , Proteína Smad2 , Fatores de Tempo , Transativadores/metabolismo , Transcrição Gênica , Ativação Transcricional , TransfecçãoRESUMO
Xylitol is transported by Streptococcus mutans via a constitutive phosphoenolpyruvate:fructose phosphotransferase system (PTS) composed of a IIABC protein. Spontaneous xylitol-resistant strains are depleted in constitutive fructose-PTS activity, exhibit additional phenotypes, and are associated with the caries-preventive properties of xylitol. Polymerase chain-reactions and chromosome walking were used to clone the fxp operon that codes for the constitutive fructose/xylitol-PTS. The operon contained three open reading frames: fxpA, which coded for a putative regulatory protein of the deoxyribose repressor (DeoR) family, fxpB, which coded for a 1-phosphofructokinase, and fxpC, which coded for a IIABC protein of the fructose-PTS family. Northern blot analysis revealed that these genes were co-transcribed into a 4.4-kb mRNA even in the absence of fructose. Inactivation of the fxpC gene conferred resistance to xylitol, confirming its function. The fxp operon is also present in the genomes of other xylitol-sensitive streptococci, which could explain their sensitivity to xylitol.
Assuntos
Proteínas de Bactérias/fisiologia , Farmacorresistência Bacteriana/genética , Genes Bacterianos , Proteínas de Membrana/fisiologia , Sistema Fosfotransferase de Açúcar do Fosfoenolpiruvato/genética , Streptococcus mutans/enzimologia , Streptococcus mutans/genética , Xilitol/farmacologia , Sequência de Aminoácidos , Proteínas de Bactérias/genética , Sequência de Bases , Passeio de Cromossomo , Proteínas de Fímbrias , Inativação Gênica , Proteínas de Membrana/genética , Dados de Sequência Molecular , Óperon/fisiologia , Reação em Cadeia da Polimerase , Transcrição Gênica , Transformação BacterianaRESUMO
Activation of transforming growth factor-beta (TGF-beta) receptors triggers phosphorylation of Smad2 and Smad3. After binding to Smad4, the complex enters the nucleus and interacts with other transcription factors to activate gene transcription. Unlike other Smads, Smad7 inhibits phosphorylation of Smad2 and Smad3, and its transcription is induced by TGF-beta, suggesting a negative feedback loop. Here, we show that TFE3 and Smad3 synergistically mediate TGF-beta-induced transcription from the Smad7 promoter by binding to an E-box and two adjacent Smad binding elements (SBEs), respectively. A precise 3-base pair spacer between one SBE and the E-box is essential. Previously, we showed that a similar arrangement between a SBE and an E-box of an element is essential for TGF-beta-dependent transcription of the plasminogen activator inhibitor-1 gene (PAI-1) and that TGF-beta-induced phosphorylation of Smad3 triggers its association with TFE3. Thus, TFE3-Smad3 response elements may represent a common target for TGF-beta-induced gene expression.