Biology of the ARL13B GTPase

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DESCRIPTION (provided by applicant): Cilia have sparked keen interest because of their proven importance in many human genetic diseases (collectively termed ciliopathies and including Joubert syndrome (JS)) and in vertebrate/mouse development; appreciation of their central role in aspects of cell signaling is growing, as well. Primary, immotile cilia are microtubule-based projections found on virtually every eukaryotic cell. We focus on the small GTPase, ARL13B, because its absence leads to defects in (1) cilia structure, (2) movement and localization of specific proteins to and (3) within cilia, and (4) patients with mutations in ARL13 have Joubert syndrome (JS), which involves renal, ocular, and brain anomalies. This application has four specific aims, in which we propose to (1) define the biochemical properties of ARL13B as a GTPase and identify and characterize novel effectors, (2) identify, purify, and characterize ARL13B GTPase activating proteins (GAPs), (3) determine the level of complexity in downstream ARL13B signaling and identify one or more effectors, (4) determine the tissue-specific consequences of JS causing ARL13B mutations in mice. We will use the complementary expertise of the two PIs to develop models for ARL13B signaling at cilia, with direct relevance to disease processes. We will pair a novel genetic deletion system in mouse embryo fibroblasts with detailed biochemical studies and modeling of ARL13B as a regulatory GTPase to develop molecular models for ARL13B's actions in cells and animals. Together, these approaches will allow us to identify and rigorously test downstream effectors and GTPase activating proteins (GAPs) of ARL13B actions in the development of the pathways affected by JS. Finally, by defining the in vivo, tissue specific function of the JS causative ARL13B mutations, we will both model the disease and identify the mechanism underlying tissue-specific Arl13b function and patient phenotypic variability. Together these aims will provide a solid experimental basis for establishing models of Arl13b actions, with clear implications for many aspects of cilia function, cell signaling, mammalian development, and human disease.