Low oxygen (O(2)) levels characterize the microenvironment of both stem cells and rapidly growing tumors. Moreover, hypoxia is associated with the maintenance of stem cell-like phenotypes and increased invasion, angiogenesis and metastasis in cancer patients. Metastatic cancers, such as breast cancer and melanoma, aberrantly express the embryonic morphogen Nodal, and the presence of this protein is correlated with metastatic disease. In this paper, we demonstrate that hypoxia induces Nodal expression in melanoma and breast cancer cells concomitant with increased cellular invasion and angiogenic phenotypes. Of note, Nodal expression remains up-regulated up to 48 h following reoxygenation. The oxygen-mediated regulation of Nodal expression occurs via a combinatorial mechanism. Within the first 24 h of exposure to low O(2), there is an increase in protein stability. This increase in stability is accompanied by an induction of transcription, mediated by the HIF-1α-dependent activation of Notch-responsive elements in the node-specific enhancer of the Nodal gene locus. Finally, Nodal expression is maintained upon reoxygenation by a canonical SMAD-dependent feed-forward mechanism. This work provides insight into the O(2)-mediated regulation of Nodal, a key stem cell-associated factor, and reveals that Nodal may be a target for the treatment and prevention of hypoxia-induced tumor progression.