Hedgehog Signaling and Cilia in Choroid Fissure Morphogenesis and Coloboma

University Of Utah, Salt Lake City UT

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Abstract

? DESCRIPTION (provided by applicant): Uveal coloboma, a condition estimated to occur in ~1:10,000 live births, is a significant cause of blindness worldwide. It is characterized by a hole or cleft in the eye, and results from defective formation or closure of the choroid fissure, a transient yet critical structure through which retinal axons exit and vasculature enters the eye. The Hedgehog (Hh) signaling pathway is vital for choroid fissure development: mutations upstream, downstream, and within the pathway can result in coloboma. Somewhat paradoxically, mutations that hyperactivate Hh signaling, as well as mutations that inactivate downstream targets, both lead to coloboma in humans, yet the specific morphogenetic defects underlying each of these models are unknown. In addition, primary cilia are required for vertebrate Hh signaling. Colobomata are associated with human ciliopathies, yet it is unclear whether the ciliopathy mutations activate or inactive Hh signaling in the eye, or how they disrupt specific morphogenetic processes to cause coloboma. Zebrafish presents an ideal model system to study this process: optical transparency and rapid development offer a unique opportunity to directly watch the choroid fissure form in vivo. We previously developed imaging and computational techniques to track and visualize cell movements throughout optic cup morphogenesis. However, choroid fissure development and the specific mechanisms disrupted in coloboma remain a mystery. In this proposal, we will determine the mechanisms underlying choroid fissure formation under normal conditions and in Hedgehog-driven zebrafish models of coloboma. I hypothesize that hyperactive Hedgehog signaling, acting through cilia, and loss of downstream effectors both result in coloboma by disrupting, in an opposing manner, cell movements underlying choroid fissure formation. Combining molecular genetics with innovative 4-dimensional live imaging and computational methods, we will test this hypothesis in the following specific aims: (1) determine how hyperactive Hh signaling, as caused by mutation in ptch2, disrupts choroid fissure formation; (2) determine how loss of the Hh downstream effector pax2 disrupts choroid fissure development to cause coloboma; and (3) determine the role of cilia signaling in choroid fissure development and ciliopathy-associated coloboma. The mechanistic experiments proposed here will define the cellular dynamics underlying normal choroid fissure formation, and the specific defects underlying three models of human coloboma, all of which affect the Hedgehog pathway: Gorlin syndrome, renal coloboma syndrome, and Joubert/COACH syndrome. Our work represents a novel strategy to understand the etiology of this potentially devastating vision disorder.