TMIE Is an Essential Component of the Mechanotransduction Machinery of Cochlear Hair Cells

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Cochlear hair cells convert sound stimuli into electrical signals by gating of mechanically sensitive ion channels in their stereociliary (hair) bundle. The molecular identity of this ion channel is still unclear, but its properties are modulated by accessory proteins. Two such proteins are transmembrane channel-like protein isoform 1 (TMC1) and tetraspan membrane protein of hair cell stereocilia (TMHS, also known as lipoma HMGIC fusion partner-like 5, LHFPL5), both thought to be integral components of the mechanotransduction machinery. Here we show that, in mice harboring an Lhfpl5 null mutation, the unitary conductance of outer hair cell mechanotransducer (MT) channels was reduced relative to wild type, and the tonotopic gradient in conductance, where channels from the cochlear base are nearly twice as conducting as those at the apex, was almost absent. The macroscopic MT current in these mutants was attenuated and the tonotopic gradient in amplitude was also lost, although the current was not completely extinguished. The consequences of Lhfpl5 mutation mirror those due to Tmc1 mutation, suggesting a part of the MT-channel conferring a large and tonotopically variable conductance is similarly disrupted in the absence of Lhfpl5 or Tmc1. Immunolabelling demonstrated TMC1 throughout the stereociliary bundles in wild type but not in Lhfpl5 mutants, implying the channel effect of Lhfpl5 mutations stems from down-regulation of TMC1. Both LHFPL5 and TMC1 were shown to interact with protocadherin-15, a component of the tip link, which applies force to the MT channel. We propose that titration of the TMC1 content of the MT channel sets the gradient in unitary conductance along the cochlea.cochlea | mechanotransducer channels | TMC1 | hair cell | LHFPL5 C ochlear hair cells detect sound stimuli by submicron vibrations of their stereociliary (hair) bundles. The stereocilia are arranged in three to four rows, stepped in height and interconnected by extracellular linkages; the most important for transduction are the tip links (1, 2), composed of cadherin-23 and protocadherin-15 (3, 4). During bundle displacements, they transmit force to activate mechanotransducer (MT) ion channels near the insertion of protocadherin-15 at the lower end of the tip link into the stereociliary tip (5, 6). The molecular identity of the pore-forming subunit of the ion channel is still controversial, but there has been a recent proposal that transmembrane channellike protein isoforms 1 and 2 (TMC1 and TMC2) (7, 8) are possible candidates (9, 10); mutations of these proteins can alter the Ca 2+ selectivity and single-channel conductance of the MT channels, implying that TMC proteins can influence ion conduction through the pore (10-12). However, in Tmc1/Tmc2 double mutants, large mechanically sensitive currents can still be evoked and flow through channels similar to native MT channels (13). Thus, an alternate view is that the TMC1 and TMC2 are accessory but not pore-forming subunits of the channel.Another likely component of the transduction mac...

Section: Discussionmentioning

confidence: 99%

Subunit determination of the conductance of hair-cell mechanotransducer channels

Beurg

Xiong

Zhao

et al. 2014

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“…Thus, biophysical attributes of the normal-polarity and unconventional channels are similar but not identical. If the channels had the same pore, such differences could reflect the presence of other subunits in the normal channel, such as LHFPL5 and TMIE (9,(32)(33)(34). It will be important to establish whether both have the same protein core or are structurally distinct, as this may provide a clue to the functional significance of the unconventional channels.…”

Section: Discussionmentioning

confidence: 99%

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...

“…It remains to be shown whether TMC1 and TMC2 can yield channel activities in heterologous expression systems (52,53), and they likely require other proteins for their function in mechanotransduction (54)(55)(56). We have attempted to ectopically express the Drosophila tmc gene product in a variety of heterologous systems.…”

Section: Differential Functions Of Different Sensory Neurons In Regulmentioning

confidence: 99%

Transmembrane channel-like ( tmc ) gene regulates Drosophila larval locomotion

Guo¹,

Wang²,

Zhang³

et al. 2016

Drosophila larval locomotion, which entails rhythmic body contractions, is controlled by sensory feedback from proprioceptors. The molecular mechanisms mediating this feedback are little understood. By using genetic knock-in and immunostaining, we found that the Drosophila melanogaster transmembrane channel-like (tmc) gene is expressed in the larval class I and class II dendritic arborization (da) neurons and bipolar dendrite (bd) neurons, both of which are known to provide sensory feedback for larval locomotion. Larvae with knockdown or loss of tmc function displayed reduced crawling speeds, increased head cast frequencies, and enhanced backward locomotion. Expressing Drosophila TMC or mammalian TMC1 and/or TMC2 in the tmc-positive neurons rescued these mutant phenotypes. Bending of the larval body activated the tmc-positive neurons, and in tmc mutants this bending response was impaired. This implicates TMC's roles in Drosophila proprioception and the sensory control of larval locomotion. It also provides evidence for a functional conservation between Drosophila and mammalian TMCs.proprioception | locomotion | mechanosensation