Critical proteins for hair cell protein trafficking and auditory function

About This Grant

PROJECT SUMMARY/ABSTRACT Hearing loss is the most common sensory deficit and often arises from dysfunction of cochlear hair cells. Hair cells express precisely localized protein complexes, including the mechanotransduction (MET) channel, which converts mechanical sound waves into electrical signals and is essential for hearing. Although this heteromeric protein complex is critical for auditory function, significant knowledge gaps remain regarding how the MET channel components are assembled, trafficked, and localized within hair cells. Proper trafficking of MET proteins—from the endoplasmic reticulum (ER) to the stereocilia via the apical cell body—is essential for hearing. Many forms of hereditary hearing loss are linked to deficits in MET protein localization, underscoring the importance of understanding the underlying molecular mechanisms. TMC1 and TMC2 are the pore-forming subunits of the MET channel, with TMC1 being the predominant form during most of life. Our recent work identified Transmembrane O-methyltransferase (TOMT) as a critical regulator of TMC1/2 localization, MET channel activity, and auditory function. TOMT is the first known molecule that regulates MET protein localization without being part of the MET complex itself. While TOMT binds TMC1, it is excluded from the stereocilia and instead localizes to the ER and cytoplasm within the hair cell body. Importantly, TOMT alone is insufficient to promote TMC1 ER exit in non-hair cells, suggesting the need for additional, hair-cell-specific factors. We identified novel molecular chaperones—PEX3, PEX16, and PEX19 (collectively, PEX proteins)—that are expressed in hair cells and interact with TOMT to enhance TMC1 stability. Mutations in PEX genes cause hearing loss and peroxisomal biogenesis disorders in humans and mice. Based on these findings, our central hypothesis is that trafficking and localization of TMC1 is regulated by a specialized, hair-cell-specific machinery involving TOMT and PEX proteins. To test this hypothesis, we will 1) define how TOMT regulates TMC1 trafficking and stereocilia localization by mapping TOMT-TMC1 interaction domains and examining the effects of deafness-linked mutations on subcellular localization and auditory function; and 2) generate and employ knock-in and conditional knockout mouse models to characterize expression patterns and manipulate inner ear gene function of PEX proteins, assessing their individual and synergistic roles with TOMT in regulating TMC1 stability, trafficking, and auditory function. This project will establish a new conceptual framework for understanding how hair cells coordinate the trafficking and localization of essential auditory proteins including TMC1. Our findings will reveal molecular checkpoints and potential therapeutic targets for hearing loss associated with defects in protein localization, including mutations in TOMT and PEX genes.