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Positional cloning of hereditary deafness genes is a direct approach to identify molecules and mechanisms underlying auditory function. Here we report a locus for dominant deafness, DFNA36, which maps to human chromosome 9q13-21 in a region overlapping the DFNB7/B11 locus for recessive deafness. We identified eight mutations in a new gene, transmembrane cochlear-expressed gene 1 (TMC1), in a DFNA36 family and eleven DFNB7/B11 families. We detected a 1.6-kb genomic deletion encompassing exon 14 of Tmc1 in the recessive deafness (dn) mouse mutant, which lacks auditory responses and has hair-cell degeneration. TMC1 and TMC2 on chromosome 20p13 are members of a gene family predicted to encode transmembrane proteins. Tmc1 mRNA is expressed in hair cells of the postnatal mouse cochlea and vestibular end organs and is required for normal function of cochlear hair cells.

The presence of one or more calcium-dependent ecto-ATPases (enzymes that hydrolyze extracellular 5′-triphosphates) in mammalian taste buds was first shown histochemically. Recent studies have established that dominant ecto-ATPases consist of enzymes now called nucleoside triphosphate diphosphohydrolases (NTPDases). Massively parallel signature sequencing (MPSS) from murine taste epithelium provided molecular evidence suggesting that NTPDase2 is the most likely member present in mouse taste papillae. Immunocytochemical and enzyme histochemical staining verified the presence of NTPDase2 associated with plasma membranes in a large number of cells within all mouse taste buds. To determine which of the three taste cell types expresses this enzyme, double label assays were performed using antisera directed against the glial glutamate/aspartate transporter, (GLAST), the transduction pathway proteins phospholipaseC β2 (PLCβ2) or the G protein subunit α-gustducin, and serotonin (5HT) as markers of type I, II or III taste cells, respectively. Analysis of the double labeled sections indicates that NTPDase2 immunoreactivity is found on cell processes that often envelop other taste cells, reminiscent of type I cells. In agreement with this observation, NTPDase2 was located to the same membrane as GLAST, indicating that this enzyme is present in type I cells. The presence of ecto-ATPase in taste buds likely reflects the importance of ATP as an intercellular signaling molecule in this system. KeywordsGustatory; ecto-ATPase; NTPDase2; taste cells; ATP signaling; mouse; taste bud Taste buds, the sensory endorgans mediating taste, reside in the epithelium of the tongue, palate and larynx. In rodents, lingual taste buds occur in three types of papillae: fungiform papillae scattered over the anterior portion of the tongue, circumvallate papillae on the posterior dorsal surface, and foliate papillae along the posterior lateral surface. Taste buds also occur at the oral entrance to the nasoincisor ducts, soft palate, epiglottis and larynx.Taste buds comprise several proliferative basal cells and 50-80 elongate epithelial cells which are heterogeneous in their morphologic and cytochemical characteristics. The differentiated taste cells in rodents are classified into three morphologic types: type I, type II, and type III as determined by cell structure, shape of the nucleus and apical processes (Murray and Murray, reviewed in Finger and Simon, 2000). These cell types usually correlate with particular histochemical features. For example, immunoreactivity for the G-protein subunit, a-gustducin, is localized to a subset of type II taste cells (Boughter et al., 1997;Yang et al., 2000b The presence of calcium-dependent ecto-ATPases (enzymes that hydrolyze extracellular 5′-triphosphates such as ATP) in mammalian taste buds has long been known as demonstrated histochemically (Iwayama and Nada, 1967;Zalewski, 1968;Iwayama, 1969;Akisaka and Oda, 1977;Barry, 1992). Since these early reports, different families of these ecto-enzymes hav...

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In the gustatory system, the recognition of sugars, amino acids and bitter-tasting compounds is the function of specialized G protein-coupled receptors. Recently, two members of novel subfamily of G protein-coupled receptors were proposed to function as taste receptors based on their speci®c expression in taste receptor cells. Here, we report the identi®cation of a third member, T1R3, of this family of receptors. T1R3 maps near the telomere of mouse chromosome 4 rendering it a candidate for the Sac locus, a primary determinant of sweet preference in mice. Consistent with its candidacy for the Sac locus, T1R3 displays taste receptor cell-speci®c expression. In addition, taster and non-taster strains of mouse harbor different alleles of T1R3.

Odorant receptors (ORs) on nasal olfactory sensory neurons are encoded by a large multigene family. Each member of the family is expressed in a small percentage of neurons that are confined to one of several spatial zones in the nose but are randomly distributed throughout that zone. This pattern of expression suggests that when the sensory neuron selects which OR gene to express it may be confined to a particular zonal gene set of several hundred OR genes but select from among the members of that set via a stochastic mechanism. Both locus-dependent and locus-independent models of OR gene choice have been proposed. To investigate the feasibility of these models, we determined the chromosomal locations of 21 OR genes expressed in four different spatial zones. We found that OR genes are clustered within multiple loci that are broadly distributed in the genome. These loci lie within paralogous chromosomal regions that appear to have arisen by duplications of large chromosomal domains followed by extensive gene duplication and divergence. Our studies show that OR genes expressed in the same zone map to numerous loci; moreover, a single locus can contain genes expressed in different zones. These findings raise the possibility that OR gene choice may be locus-independent or involve consecutive stochastic choices.

Mutations in myosin XV are responsible for congenital profound deafness DFNB3 in humans and deafness and vestibular defects in shaker 2 mice. By combining direct cDNA analyses with a comparison of 95.2 kb of genomic DNA sequence from human chromosome 17p11.2 and 88.4 kb from the homologous region on mouse chromosome 11, we have determined the genomic and mRNA structures of the human (MYO15) and mouse (Myo15) myosin XV genes. Our results indicate that full-length myosin XV transcripts contain 66 exons, are >12 kb in length, and encode 365-kDa proteins that are unique among myosins in possessing very long ϳ1200-aa Nterminal extensions preceding their conserved motor domains. The tail regions of the myosin XV proteins contain two MyTH4 domains, two regions with similarity to the membrane attachment FERM domain, and a putative SH3 domain. Northern and dot blot analyses revealed that myosin XV is expressed in the pituitary gland in both humans and mice. Myosin XV transcripts were also observed by in situ hybridization within areas corresponding to the sensory epithelia of the cochlea and vestibular systems in the developing mouse inner ear. Immunostaining of adult mouse organ of Corti revealed that myosin XV protein is concentrated within the cuticular plate and stereocilia of cochlear sensory hair cells. These results indicate a likely role for myosin XV in the formation or maintenance of the unique actin-rich structures of inner ear sensory hair cells.