Intraflagellar transport, cilia and developmental signaling

The Hedgehog (Hh) signaling pathway has crucial roles in the control of vertebrate embryonic development, stem cell maintenance and tumorigenesis. Genetic screens in the mouse first revealed the connection between mammalian Hh signaling and intraflagellar transport (IFT), required for the construction of the primary cilium. Localization of key Hh pathway components to the primary cilium in response to ligand indicate that regulated trafficking is essential for Hh signal transduction. However, the nature of the interface between the ciliogenesis pathway and the Hh pathway is not clear.;A dedicated dynein motor, cytoplasmic dynein 2, directs retrograde trafficking within the primary cilium. I show that loss of the heavy chain, Dync2h1, leads to an accumulation of IFT and Hh pathway proteins in a bloated primary cilium and a block in Shh signaling. Using combined genetic and cell biological analysis, I further define the role of Dync2h1 in neural patterning and cilia structure. Decreased anterograde trafficking dominantly suppresses the Dync2h1 phenotype: heterozygosity for a weak allele of Ift172 rescues the abnormal cilia morphology of Dync2h1 mutants, and parallels the rescue of Shh-dependent cell types in the neural tube. IFT-A proteins are also important in retrograde trafficking, but unlike Dync2h1, act as negative regulators of Shh signaling. Mutants that lack both Ift122 and Dync2h1 show ectopic pathway activation, like Ift122 single mutants. Decreased levels of Ift122 also suppresses the Dync2h1 phenotypes. These genetic interactions illustrate the cell biology underlying ciliogenesis and argue that mutations in IFT genes cause various Hh phenotypes because of their roles in cilia architecture rather than direct roles in signaling.;Mutations in the Rab family of vesicular transport protein member Rab23 cause ligand-independent activation of the Shh pathway, although no known cargos have been identified. Here I show that Rab23 mutants have short cilia and decreased Rab23 dosage can suppress the Dync2h1 phenotype like IFT complex proteins. This identifies a previously uncharacterized role for Rab23 during ciliogenesis.;Finally, I clarify the requirement of cilia in modulating cellular responses to Wnt signals and demonstrate that loss of primary cilia does not affect the response of the midgestation embryo to Wnt ligands.