The Hedgehog (Hh) signaling pathway plays crucial roles both in embryonic development and in adult stem cell function. Loss of Hh signaling during development is associated with birth defects, and inappropriate Hh pathway activation post-embryonically is associated with numerous cancer types. Despite its importance in development, physiology, and disease, our ability to therapeutically modulate Hh pathway activity is limited by an incomplete understanding of the mechanisms underlying pathway activation. To this end, a comprehensive understanding of Hh signal transduction will provide a better basis for the design and implementation of effective therapeutic approaches to diseases caused by malfunction of the Hh signaling pathway. The central mystery of Hh signal transduction is the mechanism by which the Hh receptor Patched (Ptch1 in mammals) acts to regulate the pathway activity. Mouse genetic studies have recently linked Hh signal transduction to the primary cilium, a tiny microtubule-based organelle projecting from the cell surface that senses extracellular signals and is associated with a number of human diseases collectively termed ciliopathies. Studies from several groups, including ours, show that Hh pathway components continuously traffic through the cilium. In the absence of Hh, the Ptch1 receptor localizes to the primary cilium, where it inhibits Smoothened (Smo) activation and prevents Smo accumulation in the cilia. When Hh binds to Ptch1, however, Ptch1 activity is blocked and Ptch1 leaves the cilium, leading to the accumulation of Smo in the cilium and activation of downstream signaling pathways. Ciliary enrichment of these key components of the Hh pathway, as well as their dynamic localization in response to the Hh ligand, lead us to propose that ciliary trafficking of Hh pathway proteins is crucial for Ptch1's regulation of Smo. The proposed project will elucidate the mechanism of Hh receptor function in the context of primary cilium by: (1) defining critical motifs required for Ptch1/Smo ciliary trafficking using structured illumination microscopy; (2) examining of the ciliary entry and exit kinetics of Ptch1 and Smo during Hh pathway activation with a photoconversion assay in live cells; and (3) defining the cellular factors required for Ptch1/Smo ciliary trafficking using a newly adapted biochemical (BioID) approach. Our findings will provide a paradigm for understanding how the Hh receptor functions, as well as a basis for the design and implementation of effective approaches to therapeutically modulate Hh pathway activity. Our findings will also illuminate cellular and molecular mechanisms underlying polycystic kidney disease, obesity, deafness, and other conditions associated with human ciliopathies.

Public Health Relevance

The Hedgehog (Hh) signaling pathway plays crucial roles both in embryonic development and in adult stem cell function. Loss of Hh signaling during development is associated with birth defects, whereas inappropriate Hh pathway activation post-embryonically is associated with numerous cancer types that are responsible for up to 25% of human cancer death. Our research aims to discover the central mechanism underlying how the Hh receptor travels through the primary cilium and functions as a tumor repressor, so that further research can target effective treatments or preventions of serious health issues that are caused by an Hh signaling malfunction.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM117440-03
Application #
9646360
Study Section
Molecular and Integrative Signal Transduction Study Section (MIST)
Program Officer
Gindhart, Joseph G
Project Start
2017-05-01
Project End
2022-02-28
Budget Start
2019-03-01
Budget End
2020-02-29
Support Year
3
Fiscal Year
2019
Total Cost
Indirect Cost
Name
George Washington University
Department
Anatomy/Cell Biology
Type
Schools of Medicine
DUNS #
043990498
City
Washington
State
DC
Country
United States
Zip Code
20052