Sunday, December 22, 2024
12/22/2024
The actin-based stereociliary bundle (or hair bundle) on the apex of auditory hair cells serves the critical function of converting sound energy to electric signals. Defects in hair bundle development and maintenance due to genetic and environmental factors are a leading cause for sensorineural deafness. A long-term objective of this work is to gain a mechanistic understanding of hair bundle morphogenesis programs, including secreted morphogens and their downstream signaling effectors and cytoskeletal regulators. Formation of the V-shaped hair bundle is integrally linked to planar polarization of the hair cell apical cytoskeleton, which is initiated by the peripheral migration of the kinocilium, the hair cell primary cilium. Recent advances demonstrate that hair cell intrinsic planar polarity, which we name iPCP to be concise, is regulated by several planar polarized, cell- intrinsic signaling modules and microtubule motors. At the tissue-level, the non-canonical Wnt/Planar Cell Polarity (PCP) pathway aligns hair cell orientation along the medial-lateral (or neural-abneural) axis of the cochlear duct. While disrupted tissue-level PCP signaling causes hair bundle misoriention, defective iPCP signaling results in both misoriented and misshapen hair bundles with a mispositioned kinocilium. Despite its importance for hair bundle morphogenesis, how iPCP is regulated by developmental signals and how iPCP and PCP signaling are integrated remain poorly understood. To fill these knowledge gaps, this application leverages our recent exciting discoveries and aims to elucidate the mechanisms by which a novel Wnt/G protein signaling pathway coordinately controls iPCP and PCP during hair bundle morphogenesis. While Wnt/β-catenin and Wnt/PCP pathways have been extensively studied, little is known about the in vivo function and mechanisms of Wnt/G protein signaling. Our aims are built on a strong foundation of preliminary data, showing that Wnt ligands secreted by the cochlear epithelium activate heterotrimeric G protein and PI3K signaling to control both iPCP and PCP. We have identified genes required for G protein activation in the cochlea, providing clues to the molecular mechanism. We propose to use gain- and loss-of-function alleles in vivo, ex vivo cochlear explants and cell culture studies in vitro to define critical components of the Wnt/G protein pathway and its integration with the Wnt/PCP pathway. In Aim 1, we will delineate the signal transduction machinery of the Wnt/G protein pathway by determining the roles of specific Wnt ligand, Frizzled receptors and downstream signal transducers. In Aim 2, we investigate the role of the iPCP component Par3 in Wnt-induced G protein activation. In Aim 3, we will test whether Wnt/G protein signaling is regulated by multiple G protein GEFs during cochlear morphogenesis. Ultimately, insights into this new Wnt signaling pathway may facilitate devising rational therapies to stimulate hair bundle repair following injury and to regenerate auditory hair cells through stem cell technologies.
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