Abstract
How time is measured by neural stem cells during temporal neurogenesis has remained unresolved. By combining experiments and computational modeling, we define a Shh/Gli-driven three-node timer underlying the sequential generation of motor neurons (MNs) and serotonergic neurons in the brainstem. The timer is founded on temporal decline of Gli-activator and Gli-repressor activities established through down-regulation of Gli transcription. The circuitry conforms an incoherent feed-forward loop, whereby Gli proteins not only promote expression of Phox2b and thereby MN-fate but also account for a delayed activation of a self-promoting transforming growth factor-β (Tgfβ) node triggering a fate switch by repressing Phox2b. Hysteresis and spatial averaging by diffusion of Tgfβ counteract noise and increase temporal accuracy at the population level, providing a functional rationale for the intrinsically programmed activation of extrinsic switch signals in temporal patterning. Our study defines how time is reliably encoded during the sequential specification of neurons.