Our long-term goal is to understand the etiology of polycystic kidney disease (PKD) and to provide insight for better treatment of PKD and other ciliopathies. The main goal of this project is to dissect a newly identified Arl13b-Sea-Reptin network in cilia-mediated signaling and its role in kidney cyst formation. The cilium, a previously obscure cell surface structure, plays a critical role in PKD and an increasing number of other diseases. Although many cilia-associated proteins have been identified, little is known about the functional relationship between these proteins and how signals are transduced from this cell surface organelle into the cell and the nature of elicited cellular responses. Lack of such knowledge is hindering our understanding of the role of cilia in vertebrate development and human disease. To understand cilia-mediated signaling and its role in kidney cyst formation in an integrated fashion, we combine zebrafish genetics with proteomic approaches. In a large-scale genetic screen, we isolated 13 genes that can cause kidney cyst when mutated. In addition to multiple IFT genes that are important for cilia biogenesis, also isolated in this scree are novel genes arl13b, sea and transcriptional regulators reptin and pontin. Further studies revealed that Arl13b is a ciliary protein required for cilia biogenesis, while Sea is mainly a cytoplasmic protein required for cilia signaling. Unexpectedly, we found that Arl13b co-purifies with Sea and that Sea physically interacts with Reptin through tandem affinity purification and yeast two-hybrid screens. Considering that Reptin is a well-known epigenetic/transcriptional regulator, we hypothesize that Arl13b-Sea- Reptin links the cilium to transcriptional regulation and defects in this pathway contribute to cyst formation. We propose three specific aims to test our hypothesis.
In aim 1, we will characterize the role of the Arl13b-Sea-Reptin connection in cilia-mediated signaling and PKD pathogenesis.
In aim 2, we will expand our understanding of this network by identifying and characterizing additional binding partners of Arl13b and Sea and transcriptional targets of Reptin. Finally, we will start to address the functional conservation of this network by analyzing Arl13b function in the mouse kidney. Together, this study will validate a newly identified connection between cilia on the cell surface and transcriptional regulation in the nucleus. It will also identify new players in this signaling network and thus open up new directions for future studies. Knowledge gained from this study is an important step in obtaining a detailed understanding of how cilia normally function, what their downstream targets are, and how their dysfunction contribute to PKD, thus will provide insights for rational design of effectiv treatments against this disease and other ciliopathies.
Defects in a tiny cell surface organelle called the cilium, which acts as an antenna for the cell to detect environmental signals, are responsible for a wide range of human diseases, ranging from polycystic kidney disease (PKD), obesity, to infertility and retinal degeneration. However, we still do not understand how signals from cilia are transducer into the cell and incorporated with other signals. This application proposes to study how a newly identified protein interaction network connects the cell surface cilium, transcriptional control in the nucleus and PKD pathogenesis, with the goal of providing insight for rationale design of treatments against cilia-related diseases, particularly PKD.
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