Members of the Notch family of receptors act as membrane-tethered transcription factors that are tightly associated with binary cell fate decisions. Notch signaling acts as a molecular gate that allows cells to adopt or forfeit a particular fate. Interaction of Notch with ligands triggers a sequence of proteolytic cleavages that release the intracellular domain to the nucleus; this mechanism is a target of therapies for leukemias associated with Notch activation. Although the molecular mechanism of Notch activation is well characterized, further analysis in an appropriate cellular context will provide new insight into Notch signaling.