PROJECT SUMMARY/ABSTRACT Ciliopathies are a class of inherited disorders induced by mutations of ciliary genes and manifest in dysfunction in various organs, including the olfactory system. It is believed that ciliopathy induced olfactory dysfunction is caused by defects in the multi-cilia of mature olfactory sensory neurons (mOSNs) which possess the machinery necessary for odorant detection. This proposal provides evidence that immature OSNs (iOSNs) possess primary cilia that express ARL13B, a canonical ciliary marker. ARL13B is a GTPase and a causative gene for the ciliopathy, Joubert syndrome (JS). There is a profound gap in knowledge whereby the role of ARL13B in OSNs is unknown and it is unclear if JS patients suffer from smell impairment. Intriguingly, ARL13B not only localizes to cilia but was also discovered outside of the cilium and implicated in neurite development. Few studies have explored the function of ARL13B outside of the cilium in neurons and the reports show variable effects, suggesting that this may be neuronal cell-type specific. Preliminary data in this proposal show that the deletion of ARL13B under the OMP promoter causes deformations in the structure and function of glomeruli where OSNs synapse in the olfactory bulb. Although ARL13B is widely considered a canonical marker of cilia, it is surprisingly not detected in the cilia of mOSNs which express OMP, suggesting a role of ARL13B in OSNs outside of the cilium. The central hypothesis of this proposal is that in OSNs, extraciliary ARL13B is integral for proper axon innervation of glomeruli in the olfactory bulb. The following two aims will test this hypothesis. Aim 1: Measure the effects of ARL13B loss in OSNs on axon innervation to glomeruli in the olfactory bulb. Experiments proposed in Aim 1 will use (1) adenoviral gene expression coupled with fluorescence microscopy to quantify axonal arborization and synapse formation as well as (2) electrophysiology to measure synaptic transmission from OSNs to their projection neurons in the OB of Arl13b OSN null mice. Aim 2: Elucidate the contribution of ciliary versus extraciliary ARL13B on the glomerular defects induced by the loss of ARL13B in OSNs. To test Aim 2, I will use a global knock-in mouse model with a ciliary excluded form of ARL13B. Also, adenoviral gene replacement experiments will be conducted to assess the ability of the ciliary excluded form of ARL13B to rescue the glomerular impairments in fully developed OSNs. My results will provide novel insight into how ARL13B controls axon synapse in olfactory sensory neurons and would be the first to show the penetrance of JS mutations/deletions in the olfactory system. The findings of this proposal will have broad implications for mechanisms of ciliopathy induced olfactory dysfunction, calling for the consideration of roles for ciliopathy proteins outside of the cilium in the olfactory system.