Myosin-XVa Controls Both Staircase Architecture and Diameter Gradation of Stereocilia Rows in the Auditory Hair Cell Bundles

Abstract

Mammalian hair cells develop their mechanosensory bundles through consecutive phases of stereocilia elongation, thickening, and retraction of supernumerary stereocilia. Many molecules involved in stereocilia elongation have been identified, including myosin-XVa. Significantly less is known about molecular mechanisms of stereocilia thickening and retraction. Here, we used scanning electron microscopy (SEM) to quantify postnatal changes in number and diameters of the auditory hair cell stereocilia in shaker-2 mice (Myo15^sh2) that lack both "long" and "short" isoforms of myosin-XVa, and in mice lacking only the "long" myosin-XVa isoform (Myo15^∆N). Previously, we observed large mechanotransduction current in young postnatal inner (IHC) and outer (OHC) hair cells of both these strains. Stereocilia counts showed nearly identical developmental retraction of supernumerary stereocilia in control heterozygous, Myo15^sh2/sh2, and Myo15^∆N/∆N mice, suggesting that this retraction is largely unaffected by myosin-XVa deficiency. However, myosin-XVa deficiency does affect stereocilia diameters. In control, the first (tallest) and second row stereocilia grow in diameter simultaneously. However, the third row stereocilia in IHCs grow only until postnatal day 1-2 and then become thinner. In OHCs, they also grow slower than taller stereocilia, forming a stereocilia diameter gradation within a hair bundle. The sh2 mutation disrupts this gradation and makes all stereocilia nearly identical in thickness in both IHCs and OHCs, with only subtle residual diameter differences. All Myo15^sh2/sh2 stereocilia grow postnatally including the third row, which is not a part of normal development. Serial sections with focused ion beam (FIB)-SEM confirmed that diameter changes of Myo15^sh2/sh2 IHC and OHC stereocilia resulted from corresponding changes of their actin cores. In contrast to Myo15^sh2/sh2, Myo15^∆N/∆N hair cells develop prominent stereocilia diameter gradation. Thus, besides building the staircase, the short isoform of myosin-XVa is essential for controlling the diameter of the third row stereocilia and formation of the stereocilia diameter gradation in a hair bundle.

Keywords: development; hearing; mechanotransduction; non-syndromic congenital deafness DFNB3; shaker-2 mouse.