The Notch and transforming growth factor-β (TGF-β) signaling pathways play critical roles in the control of cell fate during metazoan development. However, mechanisms of cross-talk and signal integration between the two systems are unknown. Here, we demonstrate a functional synergism between Notch and TGF-β signaling in the regulation of Hes-1, a direct target of the Notch pathway. Activation of TGF-β signaling up-regulated Hes-1 expression in vitro and in vivo. This effect was abrogated in myogenic cells by a dominant-negative form of CSL, an essential DNA-binding component of the Notch pathway. TGF-β regulated transcription from the Hes-1 promoter in a Notch-dependent manner, and the intracellular domain of Notch1 (NICD) cooperated synergistically with Smad3, an intracellular transducer of TGF-β signals, to induce the activation of synthetic promoters containing multimerized CSL- or Smad3-binding sites. NICD and Smad3 were shown to interact directly, both in vitro and in cells, in a ligand-dependent manner, and Smad3 could be recruited to CSL-binding sites on DNA in the presence of CSL and NICD. These findings indicate that Notch and TGF-β signals are integrated by direct protein–protein interactions between the signal-transducing intracellular elements from both pathways.

Nodal proteins have crucial roles in mesendoderm formation and left-right patterning during vertebrate development. The molecular mechanisms of signal transduction by Nodal and related ligands, however, are not fully understood. In this paper, we present biochemical and functional evidence that the orphan type I serine/threonine kinase receptor ALK7 acts as a receptor for mouse Nodal and Xenopus Nodal-related 1 (Xnr1). Receptor reconstitution experiments indicate that ALK7 collaborates with ActRIIB to confer responsiveness to Xnr1 and Nodal. Both receptors can independently bind Xnr1. In addition, Cripto, an extracellular protein genetically implicated in Nodal signaling, can independently interact with both Xnr1 and ALK7, and its expression greatly enhances the ability of ALK7 and ActRIIB to respond to Nodal ligands. The Activin receptor ALK4 is also able to mediate Nodal signaling but only in the presence of Cripto, with which it can also interact directly. A constitutively activated form of ALK7 mimics the mesendoderm-inducing activity of Xnr1 in Xenopus embryos, whereas a dominant-negative ALK7 specifically blocks the activities of Nodal and Xnr1 but has little effect on other related ligands. In contrast, a dominant-negative ALK4 blocks all mesoderm-inducing ligands tested, including Nodal, Xnr1, Xnr2, Xnr4, and Activin. In agreement with a role in Nodal signaling, ALK7 mRNA is localized to the ectodermal and organizer regions of Xenopus gastrula embryos and is expressed during early stages of mouse embryonic development. Therefore, our results indicate that both ALK4 and ALK7 can mediate signal transduction by Nodal proteins, although ALK7 appears to be a receptor more specifically dedicated to Nodal signaling.

Growth/differentiation factor 3 (GDF3) is highly expressed in adipose tissue, and previous overexpression experiments in mice have suggested that it may act as an adipogenic factor under conditions of high lipid load. GDF3 has been shown to signal via the activin receptor ALK4 during embryogenesis, but functional receptors in adipose tissue are unknown. In this study, we show that Gdf3 ؊/؊ mutant mice accumulate less adipose tissue than WT animals and show partial resistance to high-fat diet-induced obesity despite similar food intake. We also demonstrate that GDF3 can signal via the ALK4-homolog ALK7 and the coreceptor Cripto, both of which are expressed in adipose tissue. In agreement with a role for ALK7 in GDF3 signaling in vivo, mutant mice lacking ALK7 also showed reduced fat accumulation and partial resistance to diet-induced obesity. We propose that GDF3 regulates adiposetissue homeostasis and energy balance under nutrient overload in part by signaling through the ALK7 receptor.high-fat diet ͉ metabolism ͉ TGF-␤ ͉ energy balance ͉ insulinemia

Growth differentiation factor 11 (GDF11) contributes to regionalize the mouse embryo along its anterior-posterior axis by regulating the expression of Hox genes. The identity of the receptors that mediate GDF11 signalling during embryogenesis remains unclear. Here, we show that GDF11 can interact with type I receptors ALK4, ALK5 and ALK7, but predominantly uses ALK4 and ALK5 to activate a Smad3-dependent reporter gene. Alk5 mutant embryos showed malformations in anterior-posterior patterning, including the lack of expression of the posterior determinant Hoxc10, that resemble defects found in Gdf11-null mutants. A heterozygous mutation in Alk5, but not in Alk4 or Alk7, potentiated Gdf11 À/À -like phenotypes in vertebral, kidney and palate development in an Acvr2b À/À background, indicating a genetic interaction between the two receptor genes. Thus, the transforming growth factor-b (TGF-b) receptor ALK5, which until now has only been associated with the biological functions of TGF-b1 to TGF-b3 proteins, mediates GDF11 signalling during embryogenesis.

All major cell types in pancreatic islets express the transforming growth factor (TGF)-␤ superfamily receptor ALK7, but the physiological function of this receptor has been unknown. Mutant mice lacking ALK7 showed normal pancreas organogenesis but developed an age-dependent syndrome involving progressive hyperinsulinemia, reduced insulin sensitivity, liver steatosis, impaired glucose tolerance, and islet enlargement. Hyperinsulinemia preceded the development of any other defect, indicating that this may be one primary consequence of the lack of ALK7. In agreement with this, mutant islets showed enhanced insulin secretion under sustained glucose stimulation, indicating that ALK7 negatively regulates glucose-stimulated insulin release in ␤-cells. Glucose increased expression of ALK7 and its ligand activin B in islets, but decreased that of activin A, which does not signal through ALK7. The two activins had opposite effects on Ca 2؉ signaling in islet cells, with activin A increasing, but activin B decreasing, glucosestimulated Ca 2؉ influx. On its own, activin B had no effect on WT cells, but stimulated Ca 2؉ influx in cells lacking ALK7. In accordance with this, mutant mice lacking activin B showed hyperinsulinemia comparable with that of Alk7 ؊/؊ mice, but double mutants showed no additive effects, suggesting that ALK7 and activin B function in a common pathway to regulate insulin secretion. These findings uncover an unexpected antagonism between activins A and B in the control of Ca 2؉ signaling in ␤-cells. We propose that ALK7 plays an important role in regulating the functional plasticity of pancreatic islets, negatively affecting ␤-cell function by mediating the effects of activin B on Ca 2؉ signaling.T he signaling networks controlling metabolic processes are highly regulated and integrate the actions of both positively and negatively acting components from many different signaling pathways. Members of the transforming growth factor (TGF)-␤ superfamily, including TGF-␤s, growth and differentiation factors (GDFs), bone morphogenetic proteins (BMPs) and activins, have been implicated in the regulation of several metabolic processes. These ligands signal via distinct complexes of type I and type II receptor serine-threonine kinases, each binding to different classes of TGF-␤ ligands (1, 2). The main and most widely studied signaling pathway downstream of these receptors involves activation and nuclear translocation of Smad proteins, which in turn regulate gene transcription through multiple interactions with distinct sets of transcription factors in a cell type-specific manner (1, 2). Although less well understood, Smad-independent pathways have also been described in a variety of cell systems and involve the activation of MAP kinases, small GTPases, and Ca 2ϩ mobilization (3).Identification of cell-intrinsic factors controlling the specification and function of pancreatic endocrine cells is of major importance for understanding the regulation of blood-glucose homeostasis. The characterization of signals regul...