The internal ear's hair cells endow us with our appreciation of sound and our sense of balance. A hair cell transduces mechanical forces into electrical responses using its transduction apparatus, a molecular machine capable of detecting deflections of atomic dimensions. Residing in the mechanoreceptive organelle of a hair cell, the hair bundle, this transduction apparatus consists of a transduction channel (a mechanically gated ion channel); a tip link (an extracellular filament that transmits force to the channel's gate); and an adaptation motor (an ensemble that maintains an optimal tension in the tip link). Although the small number of hair cells in auditory and vestibular sensory epithelia has ensured that identification of molecules responsible for transduction is difficult, progress towards that goal is being made.

Our recent work has focused on three elements of the transduction apparatus, the adaptation motor, the tip link, and a plasma-membrane calcium pump. Circumstantial evidence implicates myosin Iß, an unconventional myosin isozyme, as the adaptation motor; we are presently attempting to prove that suggestion conclusively. We have designed a mutation in myosin Iß that introduces sensitivity to N⁶-modified ADP analogs, and have introduced mutated myosin Iß molecules into hair cells. If myosin Iß mediates adaptation in hair cells, the mutant myosin should be inhibited by the ADP analog. These experiments should permit us to determine conclusively whether myosin Iß does mediates adaptation.

We have also found that the transduction apparatus is more dynamic than previously thought. Broken tip links are replaced within 12 hours, and this time course suggests both that the loss of high-sensitivity hearing following loud noise exposure (e.g., rock concerts) is due to tip-link rupture and that recovery of hearing is ultimately limited by tip-link regeneration. We are continuing to characterize the process of tip-link regeneration in the expectation that clues to the link's identity can be discovered.

Both adaptation and tip-link regeneration are controlled by intracellular Ca²⁺. Hair bundles control Ca²⁺ levels through the use of mobile Ca²⁺-binding buffers and by the plasma-membrane Ca²⁺-ATPase, which exists on bundle membranes at a surprisingly high density of ~2000 per square micron. A plasma-membrane Ca²⁺-ATPase is essential for auditory and vestibular function, and we are presently investigating whether the roles suggested for the bundle Ca²⁺-ATPase — controlling Ca²⁺ that enters during transduction, signaling tip-link loss, and locally elevating extracellular Ca²⁺ levels — are the crucial roles for Ca²⁺-ATPase in the inner ear.

Future work will focus on identifying those proteins that interact with the adaptation-motor myosin and with tip links, as well as on developing strategies to identify and characterize the transduction channel. Once members of the transduction apparatus have been identified, we will study how they are assembled during development, turnover, or regeneration to make transduction so sensitive.

Gillespie, P. G., Gillespie, S. K. H., Mercer, J. A., Shah, K., and Shokat, K. M. (1999) Engineering of the myosin Iß nucleotide-binding pocket to create selective sensitivity to N⁶-modified ADP analogs. J. Biol. Chem 274, in press.

Yamoah, E. N., Lumpkin, E. A., Dumont, R. A., Smith, P. J. S., Hudspeth, A. J., and Gillespie, P. G. (1998) Plasma-membrane Ca²⁺-ATPase extrudes Ca²⁺ from hair-cell stereocilia. J. Neurosci 18:610-624.

Hasson, T., Gillespie, P. G., MacDonald, R. B., Garcia, J., Zhao, Y.-D., Yee, A. G., Mooseker, M. S., and Corey, D.P. (1997) Unconventional myosins in inner-ear sensory epithelia. J. Cell Biol. 137:1287-1307.

Zhao, Y.-D., Yamoah, E. N., and Gillespie, P. G. (1996) Regeneration of broken tip links and restoration of mechanoelectrical transduction in hair cells. Proc. Natl. Acad. USA 93:15469-15474.

Gillespie, P. G., Wagner, M. C., and Hudspeth, A. J. (1993) Identification of a 120-kD hair-bundle myosin Iß located near stereociliary tips. Neuron 11:581-594.