Mechanisms of Signaling Protein Retention in the Primary Cilium

项目总结

PROJECT SUMMARY The primary cilium is a solitary membrane extrusion from the apical surface of noncycling resting cells in mammals. It functions as a sensory organelle that is responsible for the resting cells to sense environmental conditions and communicate with adjacent cells. This organelle is composed of a microtubule-based axoneme encased in the plasma membrane, resulting in formation of a ciliary compartment that is separated from the cytoplasm by a transition zone at its base. Protein trafficking within the ciliary compartment is mediated by intraflagellar transport that transverses the compartment bi- directionally on the microtubules. Accumulating evidence suggests that the ability of the primary cilium to function as a sensory organelle depends on selective accumulation of various signaling molecules. Many components of cilium-dependent pathways are found to be transported into and out of the primary cilium in response to environmental cues. However, the mechanisms underlying the regulated trafficking remain unknown. The proposed application is to define a previously unknown mechanism in intraflagellar protein trafficking that as suggested by our preliminary data, regulates ciliary accumulation of soluble protein kinases in response to flow stress and chemical stimulation. This novel mechanism is mediated by folliculin (FLCN), a tumor suppressor, of which defects cause the Birt-Hogg-Dube (BHD) syndrome, a genetic disorder that is manifested clinically by benign tumors and cystic growth in multiple organs. In ciliated resting cells inactivation of FLCN produces profound effects on several major cilium-dependent pathways, including the Sonic Hedgehog (Shh) pathway, which becomes fully active and unresponsive to ligand stimulation. Preliminary data suggest that FLCN couples Shh ligand stimulation to ciliary accumulation and activation of Smoothened (Smo), a key but poorly understood step in Shh signaling. The application is proposed to define the underlying molecular basis and its functional significance in general ciliary signaling. Three lines of investigation will be carried out in the context of ligand and flow stress regulated Shh signaling. including: 1) determining how FLCN-mediated ciliary protein accumulation controls Smo activation in response to Shh ligand stimulation; 2) determining the role of Rab23 small GTPase in the ciliary function of FLCN; 3) determining how flow stress controls Shh signaling through FLCN. Successful completion of the proposed studies will offer a new paradigm to explain how ciliary signaling is regulated by environmental cues and elucidate the mechanism underlying the Shh ligand- induced activation of Smo.