Our research focuses on the identification and characterization of genes critical for structure and function of sensory hair cells in the cochlea and vestibule. Spontaneous mutations that cause balance dysfunction and hearing loss are ideal tools to identify genes important for the functioning of the inner ear and to elucidate their role in these sensory systems. Towards this end we concentrate our efforts on three deafness mutations: jackson circler (jc), jerker (je) and Varitint-waddler (Va). By auditory-evoked brain stem response analyses we showed that in these strains hearing impairment is completely penetrant and fully expressed in three to four week old animals. The vestibular phenotype is also fully expressed in je and Va mutants, but seems to vary in jc. To search for hearing and balance modifiers we outcrossed each of the mutations onto different genetic backgrounds. To identify their molecular identity we analyzed large segregating intersubspecific intercrosses. We constructed physical BAC contigs for each of the mutated loci and we evaluated candidate genes. The molecular cloning of these mutations should give us new insights in the development and function of the mammalian cochlea. High-resulution genetic and physical mapping of varitint-waddler identified two new members of the mucolipin gene familiy Mcoln2 and Mcoln3 in the critical interval. We found missense mutations in Mcoln3 which are responsible for deafness and pigmentaton defects in Va. Mcoln3 shows sequence and motif similarities to the transient-receptor-potential (TRP) family of ion channels. Mcoln3 localizes to cytoplasmic compartments and stereocilia of outer and inner hair cells in the organ of Corti. Based upon its motif structure and expression domain, we suggest that Mcoln3 is involved in the regulation of ion homeostasis in hair cells and melanocyte. These studies identified a new molecular link between deafness and pigmentation defects. Finally, Mcoln2 and Mcoln3 are candidate genes for hereditary and/or sporadic forms of neurosensory disorders. Work is in progress to characterize the cellular and molecular function of Mcoln3 in cochlea hair cells. We recently reported on the identification of a new allele of the protocadherin 15 gene in the mouse. Mutations in this gene cause deafness and balancing defects. This mouse model might be valuable in studying cochlea and retinal pathalogy in Usher syndrome type 1F. During the last year we concentrated our efforts on the phenotypic characterization and molecular cloning of the Jackson circler mutant. We performed extensive auditory characterization (using ABR and DPOAEs), genetic and physical mapping, and morphological developmental studies. We found that the jc mutation causes the truncation of the cochlea duct, through an arrest of duct elongation at the second turn during inner ear development. The mutation localizes to a O.2cM genetic region on proximal chromosome 10, which is approximately 170kb in physical distance. In different gentic crosses we noticed a significant suppression of auditory thresholds presumably due to the segregation of a dominant modifier locus. We are in the process of generating a congenic line and to perform a genome-wide linkage study to map this modifier. In order to understand the genetics and mechanistics of polygenic, non-mendelian inheritance of hearing loss, we surveyed outbred strains for hearing function. Outbred strains represent a genetic spectrum different from the common inbred strains and show a greater degree of allelic heterogeneity. They are models to isolate genetic interactions and to identify new deafness alleles. We found that the BlackSwiss outbred strains undergoes early-onset slow progressing hearing loss. The segregation pattern in intercrosses and backcrosses is consistent with a polygenic inheritance. Experiments are underway to map and identify the genetic variants.
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