The role of NompC (=TRPN1) for mechanotransducer gating and adaptation in the Drosophila ear

At the heart of all sensation lies a common process: The opening or closing (called the gating) of dedicated ion channels in the membranes of sensory cells. These so-called sensory transducer channels convert external stimulus energy- such as the mechanical energy contained in a sound wave- into an electrical current that flows through the sensory cell's membrane. In the case of the classical mechanical senses, i.e. the senses of touch, hearing and balance, these mechano-transducer channels are deemed to be gated in the most direct way possible, namely by the stimulus forces themselves. This direct mode of activation implies that the transducers must somehow be mechanically coupled to specialized stimulus receiver structures, such as our ear drums or the antennal sound receivers of fruit flies, for example. Somewhat ironically, however, the astonishing simplicity of their mode of activation appears to have greatly complicated the molecular identification of true mechano-transducer channels to this day. Recently, it was demonstrated that mechano-transduction in the sensory cells of the fruit-fly (Drosophila) ear, relies on mechano-transducer channels that operate according to the same biophysical principles as those in the inner ears of vertebrates. Fortunately, in Drosophila, the function of these transducer channels can be assessed in vivo, in the ears of intact flies. Given the enormous genetic tractability of the fruit fly, along with the availability of a multitude of mechano-sensory mutants, the Drosophila ear therefore constitutes an ideal system in which to probe the specific roles of identified proteins in the process of mechano-sensation, particularly their contributions to mechano-transduction. This proposal will initiate the molecular dissection of mechano-transducer function in the Drosophila ear by specifically assessing the role of an ion channel called NompC. The NompC channel, which reportedly serves mechanosensory functions in the ears of both vertebrates and invertebrates, is presently the best candidate for a true, auditory mechano-transducer channel. A common feature of mechano-transducers in the ears of both fruit flies and vertebrates seems to be their ability to adapt to a maintained stimulus: in vertebrate hair cells this adaptation is mediated by specialized adaptation motors which act to release tension from those elements that couple forces to the transducer channels, thus allowing for the channels to close despite the presence of the stimulus. Most remarkably, the adaptation of transducer channels in the Drosophila ear appears to operate in the same way as in vertebrates. Several lines of evidence have suggested an involvement of NompC in the process of mechano-transduction or mechano-transducer adaptation in Drosophila but more direct evidence remains outstanding. By using biophysical, transgenetic and modelling approaches, I will investigate the specific contribution of NompC to mechano-transduction and/or adaptation in the Drosophila ear. Despite the fact that the NompC channel, though present in the ears of non-mammalian vertebrates, seems to be absent from the ears of mammals, the study proposed here will also provide for a better understanding of our own ears' workings. Studies in non-mammalian vertebrates, such as turtles and frogs have provided much insight into fundamental mechanisms of auditory function that also apply in mammals, This study in the fruit fly is likewise expected to make a significant contribution to our molecular understanding of how ears translate the mechanical forces provided by sound into electrical signals which can be processed further on in the brain.

The major goal of this proposal is to investigate the role of the TRP channel NompC (equivalent to TRPN1) for the direct mechanical gating, and subsequent mechanical adaptation, of mechano-transducer channels (METs) in the Drosophila ear. Evidence suggests that, like the METs in vertebrate hair cells, the Drosophila transducers in hearing are formed by spring-operated, mechanically-adapting ion channels. By analyzing mechanical correlates of their gating, the function of these transducers can be probed non-invasively in the sound receivers of intact flies. NompC reportedly serves mechano-sensory function in both vertebrates and invertebrates. Previous studies in Drosophila demonstrated its involvement in (i) bristle touch sensitivity, (ii) sound-evoked CAP responses and (iii) auditory feedback amplification. All these findings are consistent with a role of NompC as a true, force-sensing mechano-transducer channel but whether NompC actually forms such a channel (or an essential component thereof) is still unknown. To this end, this study will quantify the specific contribution of NompC in mechano-transduction in the Drosophila ear by comparing the auditory systems of NompC mutant and control flies. Measurements will include (1) extracellular recording of CAP responses from the antennal nerve and (2) Laser Doppler vibrometric analyses of mechanical correlates of transducer gating and adaptation as well as (3) measurement of distortion product otoacoustic emissions (DPOAEs). In a parallel approach to NompC function, I will (4) record mechanically-evoked currents from tactile bristles of NompC mutants and controls and (5) probe their stimulus-receiving hair shafts for mechanical correlates of transducer gating.