Genome-wide linkage analyses identify Hfhl1 and Hfhl3 with frequency-specific effects on the hearing spectrum of NIH Swiss mice

BACKGROUND: The mammalian cochlea receives and analyzes sound at specific places along the cochlea coil, commonly referred to as the tonotopic map. Although much is known about the cell-level molecular defects responsible for severe hearing loss, the genetics responsible for less severe and frequency-specific hearing loss remains unclear. We recently identified quantitative trait loci (QTLs) Hfhl1 and Hfhl2 that affect high-frequency hearing loss in NIH Swiss mice. Here we used 2f1-f2 distortion product otoacoustic emissions (DPOAE) measurements to refine the hearing loss phenotype. We crossed the high frequency hearing loss (HFHL) line of NIH Swiss mice to three different inbred strains and performed linkage analysis on the DPOAE data obtained from the second-generation populations. RESULTS: We identified a QTL of moderate effect on chromosome 7 that affected 2f1-f2 emissions intensities (Hfhl1), confirming the results of our previous study that used auditory brainstem response (ABR) thresholds to identify QTLs affecting HFHL. We also identified a novel significant QTL on chromosome 9 (Hfhl3) with moderate effects on 2f1-f2 emissions intensities. By partitioning the DPOAE data into frequency subsets, we determined that Hfhl1 and Hfhl3 affect hearing primarily at frequencies above 24 kHz and 35 kHz, respectively. Furthermore, we uncovered additional QTLs with small effects on isolated portions of the DPOAE spectrum. CONCLUSIONS: This study identifies QTLs with effects that are isolated to limited portions of the frequency map. Our results support the hypothesis that frequency-specific hearing loss results from variation in gene activity along the cochlear partition and suggest a strategy for creating a map of cochlear genes that influence differences in hearing sensitivity and/or vulnerability in restricted portions of the cochlea

Phenotype and Genetics of Progressive Sensorineural Hearing Loss (Snhl1) in the LXS Set of Recombinant Inbred Strains of Mice

Progressive sensorineural hearing loss is the most common form of acquired hearing impairment in the human population. It is also highly prevalent in inbred strains of mice, providing an experimental avenue to systematically map genetic risk factors and to dissect the molecular pathways that orchestrate hearing in peripheral sensory hair cells. Therefore, we ascertained hearing function in the inbred long sleep (ILS) and inbred short sleep (ISS) strains. Using auditory-evoked brain stem response (ABR) and distortion product otoacoustic emission (DPOAE) measurements, we found that ISS mice developed a high-frequency hearing loss at twelve weeks of age that progressed to lower frequencies by 26 weeks of age in the presence of normal endocochlear potentials and unremarkable inner ear histology. ILS mice exhibited milder hearing loss, showing elevated thresholds and reduced DPOAEs at the higher frequencies by 26 weeks of age. To map the genetic variants that underlie this hearing loss we computed ABR thresholds of 63 recombinant inbred stains derived from the ISS and ILS founder strains. A single locus was linked to markers associated with ISS alleles on chromosome 10 with a highly significant logarithm of odds (LOD) score of 15.8. The 2-LOD confidence interval spans ∼4 Megabases located at position 54–60 Mb. This locus, termed sensorineural hearing loss 1 (Snhl1), accounts for approximately 82% of the phenotypic variation. In summary, this study identifies a novel hearing loss locus on chromosome 10 and attests to the prevalence and genetic heterogeneity of progressive hearing loss in common mouse strains

The Novel Mouse Mutation Oblivion Inactivates the PMCA2 Pump and Causes Progressive Hearing Loss

Progressive hearing loss is common in the human population, but we have few clues to the molecular basis. Mouse mutants with progressive hearing loss offer valuable insights, and ENU (N-ethyl-N-nitrosourea) mutagenesis is a useful way of generating models. We have characterised a new ENU-induced mouse mutant, Oblivion (allele symbol Obl), showing semi-dominant inheritance of hearing impairment. Obl/+ mutants showed increasing hearing impairment from post-natal day (P)20 to P90, and loss of auditory function was followed by a corresponding base to apex progression of hair cell degeneration. Obl/Obl mutants were small, showed severe vestibular dysfunction by 2 weeks of age, and were completely deaf from birth; sensory hair cells were completely degenerate in the basal turn of the cochlea, although hair cells appeared normal in the apex. We mapped the mutation to Chromosome 6. Mutation analysis of Atp2b2 showed a missense mutation (2630C→T) in exon 15, causing a serine to phenylalanine substitution (S877F) in transmembrane domain 6 of the PMCA2 pump, the resident Ca2+ pump of hair cell stereocilia. Transmembrane domain mutations in these pumps generally are believed to be incompatible with normal targeting of the protein to the plasma membrane. However, analyses of hair cells in cultured utricular maculae of Obl/Obl mice and of the mutant Obl pump in model cells showed that the protein was correctly targeted to the plasma membrane. Biochemical and biophysical characterisation showed that the pump had lost a significant portion of its non-stimulated Ca2+ exporting ability. These findings can explain the progressive loss of auditory function, and indicate the limits in our ability to predict mechanism from sequence alone

Genetic Analyses of the Mouse Deafness Mutations Varitint-Waddler (Va) and Jerker (Espnje)

Genetic studies on spontaneous mouse mutants with hearing defects have provided important insights into the function of genes expressed in inner ear hair cells. Here we report on our genetic analyses of the deaf mutants varitint-waddler (Va) and jerker (Espnje). A high-resolution genetic map localizes VaJ to a 0.14 ± 0.08 cM region between D3Mit85 and D3Mit259 on distal chromosome 3. By comparative mapping, the human ortholog resides at 1p22.3 between markers D1S3449 and D1S2252. To study the effect of different genetic backgrounds on the hearing phenotype, Espnje and VaJ were crossed to various inbred strains. Auditory-evoked brainstem response tests on F2 progeny demonstrate that expression, inheritance, and penetrance of the hearing phenotype are solely controlled by the mutant allele. To test for a genetic interaction between Espnje and Cdh23v, auditory function was analyzed in double heterozygotes; no significant increases of thresholds of sound pressure levels were observed. The results establish the framework for cloning the Va gene and provide valuable insights into the genetics of deafness mutations in the mouse

Mouse Chromosome 10

Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/42114/1/335-10-10-950_10n10p950.pd

Molecular characterization of TUB, TULP1, and TULP2, members of the novel tubby gene family and their possible relation to ocular diseases

Tubby, an autosomal recessive mutation, mapping to mouse chromosome 7, was recently found to be the result of a splicing defect in a novel gene with unknown function. Database searches revealed that sequences corresponding to the C terminus of the tub protein were highly conserved across a number of species including humans, mice, Caenorhabditis elegans, Arabidopsis, rice, and maize, and that tub was a member of a gene family. We describe here, TUB, the human homolog of mouse tub, and two newly characterized family members, TULP1 for tubby like protein 1 and TULP2. These three family members, which differ in the N-terminal half of the protein, share 60–90% amino acid identity across their conserved C-terminal region and have distinct tissue expression patterns. Alternatively spliced transcripts with 5′ variable sequences, three of which have been identified for the tubby gene, may mediate tissue specific expression. We also report that TUB, TULP1, and TULP2 map to human chromosomes 11p15.4, 6p21.3, and 19q13.1, respectively. TULP1 and TULP2 map within the minimal intervals identified for retinitis pigmentosa 14 on chromosome 6p21.3 and cone-rod dystrophy on chromosome 19q13.1. TULP1 and TULP2, which are expressed in the retina, make excellent candidates for these ocular diseases as a mutation within the tub gene is known to lead to early progressive retinal degeneration

Inner ear histology in ILS and ISS mice.

<p>Tolouidin blue-stained plastic sections through the cochlear duct in twelve-week-old ILS and ISS and eight-week-old C3HeB/FeJ (C3H) mice. <b>A,F,K,</b> Cross section through the organ of Corti at the mid-apical region; tm, tectorial membrane; oh, outer hair cell; ih, inner hair cell; sc, supporting cell; tC, tunnel of Corti; sM, scala media; scale bar  =  50 µm. <b>B,G,L,</b> Cross section through the spiral ligament at the mid-apical region of the cochlear duct. White arrow indicates a fibrocyte. sL, spiral ligament; scale bar  =  50 µm. <b>C,H,M,</b> Cross section through the stria vascularis at the mid-apical region of the cochlear duct. Red arrow, basal cell; white arrow, intermediate cell; green arrow, marginal cell; sV, stria vascularis; scale bar  =  10 µm. <b>D,I,N,</b> Cross section through the spiral ganglion near the base of the cochlear duct. Red arrow points to areas of degeneration. sG, spiral ganglion; scale bar  =  50 µm. <b>E,J,O,</b> Cross section through the spiral ganglion at the mid-apical region of the cochlear duct. White arrow points to a neuron and the red arrow indicates a Schwann cell. sG, spiral ganglion; scale bar  =  50 µm.</p

ABR threshold distribution in the LXS RI set.

<p><b>A–D.</b> Histograms showing threshold distributions at the click (<b>A</b>), 8- (<b>B</b>), 16- (<b>C</b>), and 32 kHz (<b>D</b>) stimuli. The Y-axis represents the number of RI strains, and the X-axis denotes the threshold level. Histograms were fitted with a normal Gaussian distribution. r²  =  goodness-of-fit.</p

Auditory characteristics of ILS and ISS mice.

<p><b>A.</b> ABR thresholds at click (c) and pure tone pips at 8-, 16-, and 32 kHz at twelve and 26 weeks (wks) of age. Data are given as mean ± SD. <b>B.</b> Wave I ABR amplitudes at 8- and 16 kHz stimuli at 60 dB SPL input levels. Data are given as mean ± SD. µV, microvolt. <b>C.</b> Latencies of ABR waves I – V at a 16 kHz stimulus of 60 dB SPL. Data are given as mean ± SD; msec, milliseconds. <b>D.</b> DPOAE output levels at 2f1-f2 in dB SPL over f2 frequency range 6–56 kHz (left panel). The right panel shows I/O function at f2 = 16 kHz. Data are given as mean ± SEM. <b>E.</b> Endocochlear potentials. Data are given as mean ± SD; mV, milliVolt. For all panels: C3HeB/FeJ (blue), ILS (green) and ISS (red); **<i>p</i><0.01; ***<i>p</i><0.001.</p