Single-molecule force spectroscopy reveals the dynamic strength of the hair-cell tip-link connection

The conversion of auditory and vestibular stimuli into electrical signals is initiated by force transmitted to a mechanotransduction channel through the tip link, a double stranded protein filament held together by two adhesion bonds in the middle. Although thought to form a relatively static structure, the dynamics of the tip-link connection has not been measured. Here, we biophysically characterize the strength of the tip-link connection at single-molecule resolution. We show that a single tip-link bond is more mechanically stable relative to classic cadherins, and our data indicate that the double stranded tip-link connection is stabilized by single strand rebinding facilitated by strong cis-dimerization domains. The measured lifetime of seconds suggests the tip-link is far more dynamic than previously thought. We also show how Ca²⁺ alters tip-link lifetime through elastic modulation and reveal the mechanical phenotype of a hereditary deafness mutation. Together, these data show how the tip link is likely to function during mechanical stimuli.