PROJECT SUMMARY
The formation of complex tissue and organs systems in developing organisms depends on the ability of
dividing stem and progenitor cells to properly integrate extracellular signals present in the embryonic
environment. The combined actions of these signals in turn direct diverse processes such as progenitor
maintenance, differentiation, and assignment of specific cell fates. A key step towards understanding the basis
of and means for preventing birth defect thus lies in defining how different signaling pathways mechanistically
intersect, permitting the activation of one signaling pathway to influence responses to other signals. Moreover,
the combinatorial activities have the potential to extend the range of outcomes possible from a limited range of
developmental signals. Our studies in the developing spinal cord have identified an unexpected role for Notch
receptor signaling in modulating the response of cells to the tissue morphogen Sonic hedgehog (Shh). Both
activation and inactivation of the Notch pathway alters the dorsoventral register of neural progenitors, leading
to corresponding changes in neuronal and glial cell fates (Kong et al. Dev Cell, 2015). In tracking down the
mechanism behind these effects, we discovered that Notch signaling influences the trafficking of the Shh
receptor Patched1 (Ptch1) and the key downstream Shh effector Smoothened (Smo) to primary cilia, leading to
changes in downstream Shh pathway activities. Importantly, this role for Notch can be seen in multiple cell
types including mouse and human neural progenitors, fibroblasts, and skeletal myoblasts, suggesting it may be
a general feature of mammalian cells. Collectively, our studies reveal a novel and surprisingly proximal role for
Notch in shaping the interpretation of the Shh morphogen gradient and thereby impacting cell fate
determination. The means by which Notch influences the trafficking of Shh signaling proteins, however,
remains unknown. Our preliminary data suggest that these actions of Notch are most likely transcriptionally
mediated, raising the questions of what are the target genes regulated by Notch, and how do they impact the
trafficking of Shh pathway components, and possibly other signaling proteins, to primary cilia? Moreover, do
defects in this Notch-mediated pathway contribute to congenital defects affecting ciliary transport, collectively
termed ciliopathies? Our proposed studies will address these questions, first by identifying the transcriptional
targets of Notch and its downstream effector Hes1 that coincide with changes in ciliary trafficking, and second
by developing a platform for investigating the function of these newly identified Notch target genes in Shh
signaling through CRISPR/Cas9-mediated deletions. With this approach, we will: a) reveal the nature of the
functional intersection between the Notch and Shh transduction pathways in neural fate selection and b)
identify new regulators of Shh signaling and protein trafficking to primary cilia more generally.
