Hair-Bundle Lipids

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DESCRIPTION (provided by applicant): The goal of this proposal is to determine the role of lipids in mechanotransduction by hair cells. We propose three specific aims. In Aim 1, we will determine which lipids are present in hair bundles using mass spectrometry and histochemical localization. In addition, we will measure the turnover of polyphosphatidylinositols and determine the identity, localization, and activities of enzymes that synthesize and degrade them. These experiments are shaped by the hypothesis that hair bundles have an unusual lip d composition and distribution and that these characteristics are important for transduction. In Aim 2, using fluorescence recovery after photobleaching, we will determine dynamics of the bulk hair-bundle membrane as well as the dynamics of individual lipid species. These experiments are motivated by the hypothesis that bundle membranes may have domains of reduced lipid mobility. Finally, in Aim 3 we will determine how lipids regulate transduction and adaptation. These experiments will test the hypotheses that (a) membrane association regulates myosin-1c activity, (b) myosin-1c binding to membranes alters lipid dynamics, (c) the adaptation motor and transduction channel interact in a specialized lipid domain, and (d) that the elasticity measured in the "gating spring" is due to properties of the stereocilia membrane itself. Together, the experiments in this proposal will allow us to determine how lipids regulate transduction and adaptation.