A Role for the Calcium activated Chloride Channel TMEM16a in Primary Ciliogenesis

项目总结

PROJECT SUMMARY The primary cilium, an apical cellular appendage akin to a cellular antenna, is involved in a variety of physiological processes including development, proliferation, and cell survival. Mutations in many genes linked to ciliary proteins cause a family of human diseases termed ciliopathies which include Joubert Syndrome and Bardet- Biedl syndrome. Evidencing the ubiquitous function of the primary cilium, these diseases present diverse clinical phenotypes affecting many of the body’s organ systems. As fundamental signaling hubs with distinct protein and lipid compositions relative to the rest of the cell, primary cilia are highly regulated in both their formation and function. Ion channels represent a unique class of proteins which regulate signaling events in many cellular processes, but are only beginning to be understood and appreciated in the context of the cilium. We are using a combination of electrophysiology and cell imaging to study the role of the calcium activated chloride channel ANO1 in regulating primary ciliogenesis and ciliary signaling. We recently found that Ano1 knock out results in fewer and shorter cilia compared to cells expressing ANO1. Furthermore, knock out of Ano1 in mice results in neonatal lethality due to tissue malformation mirroring classical ciliopathy hallmarks. We also recently characterized a novel cellular structure, the nimbus, which contains ANO1 and appears spatio-temporally related to ciliogenesis. We will test the hypothesis that ANO1 plays a role in regulating cilium formation and function. First, we will characterize the localization and activity of ANO1 during ciliogenesis. These experiments when during ciliogenesis ANO1 activity is important, and what step(s) of ciliogenesis ANO1 regulates. Finally, we will quantify ciliary signaling outputs in the presence and absence of ANO1. This work will allow us to elucidate the role of ANO1 in primary ciliogenesis, the functional consequence of loss of ANO1, and provide insight into primary cilium ion channel biology overall. Understanding the molecular pathways will provide valuable insights into the study of calcium activated chloride channels, and ion channels in the primary cilium. This fundamental knowledge can then be applied to primary cilia in the context of organismal development and ciliopathies and may provide new insight into pathways and targets for the treatment of these diseases.