During embryonic development TGFbeta signalling is essential for gastrulation and patterning, including specification of body axes. In the adult it plays roles in tissue homeostasis by promoting differentiation and counteracting proliferation, it also exhibits tumour-suppressing functions in several tissues. Deregulation of signalling is implicated in various diseases, including vascular disorders, fibrosis and cancer. TGFbeta regulates epithelial to mesenchymal transition (EMT), a migratory cellular behaviour required during gastrulation and organogenesis, while within tumours EMT promotes metastasis. The TGFbeta ligand Nodal is essential for gastrulation and anterior-posterior (A-P) patterning of the vertebrate embryo. A-P axis specification is the best-characterised dose-dependent function of this pathway, with high levels required for anterior and lower for more posterior structures. However, it is not known how this is established. Several regulatory factors have been identified suggesting that TGFb/Nodal signalling levels are tightly regulated. In mice, loss of function mutations in single intracellular negative regulators of the pathway, such as SnoN, Ski, Smad7 and Smad6, failed to impair A-P patterning raising concerns about their normal function in vivo. On the contrary, loss of Arkadia/RNF111, which has been shown to degrade all the above negative factors to enhance intracellular Nodal signalling, causes severe anterior truncation of the A-P embryonic axis. This implies that in its absence there is hyperactivity of negative regulators that reduces signalling. We propose to examine this using mouse genetics and A-P patterning in the embryo, along with the differentiation of embryonic stem cells towards anterior endoderm-foregut cell fates in culture. We expect our results to shed light on the dose-dependent functions of Nodal by revealing the importance of intracellular regulation in establishing the different responses and the underlying mechanisms.