Transforming growth factor–β (TGF-β) superfamily members regulate a plethora of developmental processes, and disruption of their activity has been implicated in a variety of human diseases ranging from cancer to chondrodysplasias and pulmonary hypertension. Intense investigations have revealed that SMAD proteins constitute the basic components of the core intracellular signaling cascade and that SMADs function by carrying signals from the cell surface directly to the nucleus. Recent insights have revealed how SMAD proteins themselves are regulated and how appropriate subcellular localization of SMADs and TGF-β transmembrane receptors is controlled. Current research efforts investigating the contribution of SMAD-independent pathways promise to reveal advances to enhance our understanding of the signaling cascade.

Myostatin, a transforming growth factor β (TGF-β) 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 (TβRI), to induce phosphorylation of Smad2/Smad3 and activate a TGF-β-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.

The transforming growth factor-β (TGFβ) and Wnt/wingless pathways play pivotal roles in tissue specification during development. Activation of Smads, the effectors of TGFβ superfamily signals, results in Smad translocation from the cytoplasm into the nucleus where they act as transcriptional comodulators to regulate target gene expression. Wnt/wingless signals are mediated by the DNA-binding HMG box transcription factors lymphoid enhancer binding factor 1/T cell-specific factor (LEF1/TCF) and their coactivator β-catenin. Herein, we show that Smad3 physically interacts with the HMG box domain of LEF1 and that TGFβ and Wnt pathways synergize to activate transcription of the Xenopus homeobox gene twin ( Xtwn ). Disruption of specific Smad and LEF1/TCF DNA-binding sites in the promoter abrogates synergistic activation of the promoter. Consistent with this observation, introduction of Smad sites into a TGFβ-insensitive LEF1/TCF target gene confers cooperative TGFβ and Wnt responsiveness to the promoter. Furthermore, we demonstrate that TGFβ-dependent activation of LEF1/TCF target genes can occur in the absence of β-catenin binding to LEF1/TCF and requires both Smad and LEF1/TCF DNA-binding sites in the Xtwn promoter. Thus, our results show that TGFβ and Wnt signaling pathways can independently or cooperatively regulate LEF1/TCF target genes and suggest a model for how these pathways can synergistically activate target genes.