The role of phosphoinositides in the primary cilium length and stability

Primary cilia are sensory organelles on the surface of many postmitotic cells. Several aspects of ciliary function have remained incompletely understood, including the observation that the ciliary membrane is continuous with the plasma membrane but differs in its phospholipid composition. Specifically, the density of the signalling lipid phosphatidylinositol 4,5-bisphosphate (PIP2) is normal in the ciliary base but much reduced in the ciliary tip. In order to determine the functional significance of this distribution, chemically induced protein dimerization was used to selectively synthesize or degrade PIP2 in the ciliary membrane. Using life cell microscopy, ciliary fission was observed when PIP2 was synthesized locally in the cilium. Ciliary fission was dependent on actin polymerisation in the cilium, on the Rho kinase Rac, on aurora kinase A (AurkA) and on histone deacetylase 6 (HDAC6). The phenotype and signalling pathway of ciliary fission was thus similar to ciliary decapitation required for serum- induced cell cycle re-entry. In addition, ciliary fission was reminiscent of vesicle budding induced by agonist binding when retrograde transport or endocytosis are blocked. I found that agonist binding increased ciliary PIP2 and that vesicle budding requires ciliary PIP2, indicating that the signalling pathways of fission and vesicle budding overlap. Ciliopathies are a group of human diseases characterized by ciliary dysfunction. Mutation in the PI 5-phosphatase Inpp5e can cause ciliopathies. In order to relate my findings to this disease I knocked down Inpp5e in our expression system. The resulting phenotype was blocked by reducing ciliary PIP2, and by inhibiting actin polymerisation, AurkA or HDAC6. These findings suggest that a programme comprising PIP2 increase, actin polymerisation, Rac, AurkA and HDAC is used for diverse cellular events. This programme is abnormally activated in Inpp5e-induced ciliopathies and can be blocked by targeting AurkA or HDAC6. When ciliary PIP2 was depleted, I observed a growing ciliary length, which was dependent on exocytosis of Golgi-derived vesicles. Since endocytosis requires PIP2, this finding is consistent with the hypothesis that the presence of PIP2 in the ciliary base allows membrane turnover by exocytosis and endocytosis. Taken together, ciliary PIP2 appears to be a key regulator of ciliary stability, length and membrane turnover