Stepwise Morphological and Functional Maturation of Mechanotransduction in Rat Outer Hair Cells

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178

The mechanotransducer channels of auditory hair cells are gated by tip-links, oblique filaments that interconnect the stereocilia of the hair bundle. Tip-links stretch from the tips of stereocilia in the short and middle rows to the sides of neighboring, taller stereocilia. They are made of cadherin-23 and protocadherin-15, products of the Usher syndrome type 1 genes USH1D and USH1F, respectively. In this study we address the role of sans, a putative scaffold protein and product of the USH1G gene. In Ush1g −/− mice, the cohesion of stereocilia is disrupted, and both the amplitude and the sensitivity of the transduction currents are reduced. In Ush1g fl/fl Myo15-cre +/− mice, the loss of sans occurs postnatally and the stereocilia remain cohesive. In these mice, there is a decrease in the amplitude of the total transducer current with no loss in sensitivity, and the tips of the stereocilia in the short and middle rows lose their prolate shape, features that can be attributed to the loss of tip-links. Furthermore, stereocilia from these rows undergo a dramatic reduction in length, suggesting that the mechanotransduction machinery has a positive effect on F-actin polymerization. Sans interacts with the cytoplasmic domains of cadherin-23 and protocadherin-15 in vitro and is absent from the hair bundle in mice defective for either of the two cadherins. Because sans localizes mainly to the tips of short-and middle-row stereocilia in vivo, we conclude that it belongs to a molecular complex at the lower end of the tip-link and plays a critical role in the maintenance of this link.auditory mechanoelectrical transduction | Usher syndrome type 1 | deafness | conditional knockout mice | organ of Corti

Section: Resultsmentioning

confidence: 99%

Usher type 1G protein sans is a critical component of the tip-link complex, a structure controlling actin polymerization in stereocilia

Caberlotto

Michel

Foucher

et al. 2011

128

178

“…A difficulty with assigning significance to the reverse-polarity MT currents stems from them being recorded only under abnormal circumstances, whether by tip-link disruption or mutations that adversely affect mechanotransduction. Previous work had, however, suggested that this type of current may also be present constitutively during hair-cell development (10,12,14,18). More detailed investigation showed that hair cells first displayed mechanical sensitivity around birth, when it was manifested as a reverse-polarity current, always preceding the normal-polarity current ( Fig.…”

Section: Significancementioning

confidence: 88%

Development and localization of reverse-polarity mechanotransducer channels in cochlear hair cells

Beurg

Goldring

Ricci

et al. 2016

Cochlear hair cells normally detect positive deflections of their hair bundles, rotating toward their tallest edge, which opens mechanotransducer (MT) channels by increased tension in interciliary tip links. After tip-link destruction, the normal polarity of MT current is replaced by a mechanically sensitive current evoked by negative bundle deflections. The "reverse-polarity" current was investigated in cochlear hair cells after tip-link destruction with BAPTA, in transmembrane channel-like protein isoforms 1/2 (Tmc1:Tmc2) double mutants, and during perinatal development. This current is a natural adjunct of embryonic development, present in all wild-type hair cells but declining after birth with emergence of the normal-polarity current. Evidence indicated the reverse-polarity current seen developmentally was a manifestation of the same ion channel as that evident under abnormal conditions in Tmc mutants or after tip-link destruction. In all cases, sinusoidal fluid-jet stimuli from different orientations suggested the underlying channels were opened not directly by deflections of the hair bundle but by deformation of the apical plasma membrane. Cell-attached patch recording on the hair-cell apical membrane revealed, after BAPTA treatment or during perinatal development, 90-pS stretch-activated cation channels that could be blocked by Ca 2+ and by FM1-43. High-speed Ca 2+ imaging, using swept-field confocal microscopy, showed the Ca 2+ influx through the reverse-polarity channels was not localized to the hair bundle, but distributed across the apical plasma membrane. These reverse-polarity channels, which we propose to be renamed "unconventional" mechanically sensitive channels, have some properties similar to the normal MT channels, but the relationship between the two types is still not well defined.hair cells | mechanotransducer channels | calcium imaging | cochlea | transmembrane channel-like protein I on channels sensitive to mechanical deformation of the cell membrane are widely distributed in vertebrates and are integral to the function of specialized mechanoreceptors, such as those in the sensory neurons of the skin, vasculature, or inner ear. The molecular structure of these mechanically gated channels has been the subject of much recent research and speculation (1-4) because they are the last category of vertebrate ion-channel (after voltage-gated and ligand-activated channels) evading characterization. Although success was achieved by assigning piezo2 as a transduction channel in many cutaneous touch receptors (3), uncertainty persists about the composition of the mechanotransducer (MT) channel in hair cells of the inner ear (2, 4). There, the MT channels reside in the stereociliary (hair) bundle, where they are activated by deflections of the bundle toward the taller edge of the staircase, and so increasing tension in the oblique extracellular tip links connecting adjacent stereocilia. Transmembrane channel-like protein isoforms 1 and 2, TMC1 and TMC2 (5), have been suggested as candidates for the ha...

“…ENU Mutagenesis, Functional Studies, Positional Cloning, Histology, and Biochemistry. ENU mutagenesis, analysis of ABRs and distortion product otoacoustic emissions, test of vestibular function, staining of sections and whole mounts, electron microscopy, positional cloning, the expression and purification of recombinant CDH23 and PCDH15, and protein interaction studies have been described previously (16,35,47,48). Tip tenting was quantified by counting tented stereocilia in the medium row in inner hair cell bundles (in 2-6 hair cells per time point and genotype).…”

Section: Methodsmentioning

confidence: 99%

A mouse model for nonsyndromic deafness (DFNB12) links hearing loss to defects in tip links of mechanosensory hair cells

Schwander

Xiong

Tokita

et al. 2009

Deafness is the most common form of sensory impairment in humans and is frequently caused by single gene mutations. Interestingly, different mutations in a gene can cause syndromic and nonsyndromic forms of deafness, as well as progressive and age-related hearing loss. We provide here an explanation for the phenotypic variability associated with mutations in the cadherin 23 gene (CDH23). CDH23 null alleles cause deaf-blindness (Usher syndrome type 1D; USH1D), whereas missense mutations cause nonsyndromic deafness (DFNB12). In a forward genetic screen, we have identified salsa mice, which suffer from hearing loss due to a Cdh23 missense mutation modeling DFNB12. In contrast to waltzer mice, which carry a CDH23 null allele mimicking USH1D, hair cell development is unaffected in salsa mice. Instead, tip links, which are thought to gate mechanotransduction channels in hair cells, are progressively lost. Our findings suggest that DFNB12 belongs to a new class of disorder that is caused by defects in tip links. We propose that mutations in other genes that cause USH1 and nonsyndromic deafness may also have distinct effects on hair cell development and function.cadherin 23 ͉ Cdh23 ͉ Usher syndrome ͉ progressive hearing loss