Project Summary: In the vestibular system of the inner ear, motion is detected via the mechanical deflection of a bundle of stereocilia located at the top of sensory receptor hair cells. The bundle is morphologically and physiologically polarized because only movements of the bundle towards a lone kinocilium positioned at one side of the apical cell surface are able to produce excitatory responses to acceleration or gravity. Thus, the range of motion that can be detected by an individual hair cell is determined by the polarized orientation of the stereociliary bundle. The utricle and saccule contain thousands of vestibular hair cells that detect linear accelerations which are divided between two groups that have opposite stereociliary bundle orientations and therefore respond to motion in opposite directions. These two groups of hair cells meet at a cellular junction that anatomically divides the utricle and saccule called the Line of Polarity Reversal (LPR). Our goal is to identify the cellular and molecular mechanisms that direct the development of planar polarity and the LPR, and underlie the formation of a sensory representation of gravity and acceleration in the mouse utricle and saccule. This will be addressed throughout the course of the project using a combination of knockout and transgenic mouse models, molecular genetic approaches to evaluate the transcriptional regulation of an essential regulatory kinase called STK32A, and biochemical and proteomics approaches to identify STK32A substrates. Specifically, we will test the hypothesis that patterns of Stk32a gene expression are determined by the transcription factor EMX2, and that STK32A guides stereociliary bundle orientation in hair cells that do not express EMX2. STK32A is a dark kinase with previously unknown function, which my lab has recently shown to be necessary and sufficient to determine stereociliary bundle orientation. This effort is significant and innovative because it introduces post-translational modification by kinase signaling as a new regulatory mechanism guiding hair cell planar polarity, and because it focuses on the EMX2-negative hair cells that were previously excluded from studies, including my own, that focused specifically on EMX2. Although directed towards the development of vestibular planar polarity, we anticipate that this research will impact our understanding of auditory planar polarity as well as other organ systems that rely upon cellular polarization for growth or function.