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Author ORCID Identifier

https://orcid.org/0000-0002-8422-9245

Date Available

6-25-2027

Year of Publication

2026

Document Type

Doctoral Dissertation

Degree Name

Doctor of Philosophy (PhD)

College

Medicine

Department/School/Program

Physiology

Faculty

A. Catalina Velez Ortega

Abstract

Auditory hair cells detect sound through highly organized, staircase-shaped bundles of actin protrusions known as stereocilia. The mechanical deflection of the hair bundle increases the tension in extracellular connectors known as tip links. This leads to the gating of mechano-electrical transduction (MET) channels at stereocilia tips of the second and third row, thus converting mechanical stimuli into electrical signals. Even at rest, a small MET channel current permits constitutive cation influx, which has been implicated in regulating the stereocilia cytoskeleton structure of the shorter rows of stereocilia (transducing rows). In mammals, auditory hair cells do not regenerate, requiring the bundle to maintain structural integrity and precise geometry for decades. However, the molecular mechanisms that couple mechanotransduction activity to long-term cytoskeletal and tip link maintenance remain poorly understood.

In the first part of this dissertation, I investigate the role of MYO15A isoforms in activity-dependent cytoskeleton remodeling of stereocilia. MYO15A is a non-conventional myosin, essential for stereocilia elongation, bundle formation, and maintenance of stereocilia in mature hair cells. Defects in MYO15A lead to profound hearing loss and abnormally short stereocilia. Three isoforms of MYO15A have been identified in the auditory hair cells. MYO15A-1 traffics to the second and third rows of stereocilia. MYO15A-2 is preferentially trafficked to the tips of stereocilia from the tallest row of the developing bundle. Expression of MYO15A-3 increases in mature hair cells as the developmental MYO15A-2 fades away. Using different mouse models and microscopy techniques, I demonstrate that MYO15A isoforms contribute to MET-dependent remodeling of the stereocilia actin core. Hair cells lacking functional MYO15A (all isoforms) did not exhibit MET-dependent remodeling in their stereocilia cytoskeleton. In contrast, hair cells lacking only MYO15A-1 showed exaggerated MET-dependent stereocilia remodeling, including remodeling in stereocilia from the tallest ‘non-transducing’ row of the bundle. We conclude that MYO15A isoforms enable and fine-tune the MET-dependent remodeling of the actin cytoskeleton in transducing stereocilia, while also contributing to the stability and identity of the tallest row of the bundle.

In the second part, I examine how hair cells preserve their tip links and hence their mechanotransduction function during exaggerated MET-dependent stereocilia remodeling. Tip links are essential for mechanotransduction, as they couple mechanical stimuli to the opening of MET channels at the tips of shorter stereocilia. The absence of tip link proteins can cause deafness and severe morphological changes to the bundle. Here, I demonstrate that tip link breakage and repair can occur during MET-dependent remodeling of the stereocilium cytoskeleton. Following short-term blockage of MET channels, stereocilia in the transducing rows undergo progressive shortening accompanied by tip link loss, altered length of remaining tip links, and reduced MET currents. Also, the upper tip link anchoring and presumed tension-maintaining site known as the upper tip link density (UTLD) remains near its original position and becomes uncoupled from the shortened stereocilia. At later stages, tip link number, apparent length, and MET currents are restored. Strikingly, the UTLD reemerges at a new position aligned with the remodeled height of the transducing stereocilia, indicating plastic relocation of the upper tip link insertion site to reestablish the normal tip link configuration.

Altogether, this work demonstrates that mechanotransduction is not only a sensory process but also a regulator of hair bundle architecture. From MYO15A-dependent cytoskeletal remodeling to dynamic repositioning of the tip link and its intracellular anchoring, hair cells dynamically preserve structural integrity of the bundle and the MET tensioning machinery during stereocilia remodeling.

Digital Object Identifier (DOI)

https://doi.org/10.13023/etd.2026.329

Archival?

Archival

Funding Information

This work was supported by the University of Kentucky College of Medicine and Department of Physiology funds to A. Catalina Velez Ortega,National Institutes of Health  R01DC021325 and R21DC017247 to A. Catalina Velez Ortega, the University of Kentucky College of Medicine Graduate Student Dissertation Research Grant to Ana Lopez- Porras, and National Institutes of Health R01DC014658 and R01DC019054 to Gregory Frolenkov. 

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