A novel splice site mutation of myosin VI in mice leads to stereociliary fusion caused by disruption of actin networks in the apical region of inner ear hair cells

Abstract

An unconventional myosin encoded by the myosin VI gene (MYO6) contributes to hearing loss in humans. Homozygous mutations of MYO6 result in nonsyndromic profound congenital hearing loss, DFNB37. Kumamoto shaker/waltzer (ksv) mice harbor spontaneous mutations, and homozygous mutants exhibit congenital defects in balance and hearing caused by fusion of the stereocilia. We identified a Myo6c.1381G>A mutation that was found to be a p.E461K mutation leading to alternative splicing errors in Myo6 mRNA in ksv mutants. An analysis of the mRNA and protein expression in animals harboring this mutation suggested that most of the abnormal alternatively spliced isoforms of MYO6 are degraded in ksv mice. In the hair cells of ksv/ksv homozygotes, the MYO6 protein levels were significantly decreased in the cytoplasm, including in the cuticular plates. MYO6 and stereociliary taper-specific proteins were mislocalized along the entire length of the stereocilia of ksv/ksv mice, thus suggesting that MYO6 attached to taper-specific proteins at the stereociliary base. Histological analysis of the cochlear hair cells showed that the stereociliary fusion in the ksv/ksv mutants, developed through fusion between stereociliary bundles, raised cuticular plate membranes in the cochlear hair cells and resulted in incorporation of the bundles into the sheaths of the cuticular plates. Interestingly, the expression of the stereociliary rootlet-specific TRIO and F-actin binding protein (TRIOBP) was altered in ksv/ksv mice. The abnormal expression of TRIOBP suggested that the rootlets in the hair cells of ksv/ksv mice had excessive growth. Hence, these data indicated that decreased MYO6 levels in ksv/ksv mutants disrupt actin networks in the apical region of hair cells, thereby maintaining the normal structure of the cuticular plates and rootlets, and additionally provided a cellular basis for stereociliary fusion in Myo6 mutants.

ELMOD1 stimulates ARF6-GTP hydrolysis to stabilize apical structures in developing vestibular hair cells

“…For a different example in hair cells, the Usher syndrome type I deafness mutants all show disorganized stereocilia, consistent with the suggestion that the Usher proteins are responsible for constructing interstereocilia linkages (Cosgrove and Zallocchi, 2014). Another common phenotype seen in hair cells is shared by Myo6, Ptprq, and Rdx mutants, which all have protruding apical membranes and eventual stereocilia elongation and fusion (Self et al, 1999;Kitajiri et al, 2004;Goodyear et al, 2012;Seki et al, 2017); a similar phenotype is seen with conditional Cdc42 mutants (Ueyama et al, 2014). Utricle hair cells of rda/rda mice clearly share this phenotype, as do cochlear hair cells (Johnson et al, 2012).…”

OHC-TRECK: A Novel System Using a Mouse Model for Investigation of the Molecular Mechanisms Associated with Outer Hair Cell Death in the Inner Ear

“…2a). By staining for myosin VI (MYO6), a known hair cell marker 18,29 , we confirmed that OHCs from Prestin - hDTR mice were almost all lost from the organ of Corti 7 days after administration of DT. Strong MYO6 signals were detected in the cuticular plate and the cytoplasm in both IHCs and OHCs from the apical, middle, and basal areas of the cochleae of DT-treated wild-type mice (Figs 2b and S2), suggesting that DT administration does not affect the hair cells of wild-type mice.…”

Genomic non-redundancy of the mir-183/96/182 cluster and its requirement for hair cell maintenance

“…This suggests that one or more targets regulated by miR-183 and miR-96 but not miR-182 are crucial for normal hair cell function. Again using CSmirTar, we filtered predicted miR-183/96/182 target genes with the disease term “sensorineural hearing loss,” and found two genes that meet these criteria, FGFR1 , which may regulate patterning of the developing cochlear duct 63 , and MYO6 , which is expressed in mature hair cells and is critical for the integrity of stereocilia 64 . Further insight into the biology of these miRNAs will be gained by comprehensively examining their potential targets and identifying additional common genes and pathways that they may co-regulate.…”