Jeffrey D. Ohmen

Positional cloning of the combined hyperlipidemia gene Hyplip1.

We have developed technologies that simplify genomic library construction and screening, substantially reducing both the time and the cost associated with traditional library screening methods and facilitating the generation of gene-targeting constructs. By taking advantage of homologous recombination in Escherichia coli, we were able to use as little as 80 bp of total sequence homology to screen for a specific gene from a genomic library in plasmid or phage form. This method, called recombination cloning (REC), takes only a few days instead of the several weeks required for traditional plaque-lift methods. In addition, because every clone in the mouse genomic library we have constructed has a negative selection marker adjacent to the genomic insert, REC screening can generate gene-targeting vectors in one step, from library screening to finished construct. Conditional targeting constructs can be generated easily with minimal additional manipulation.

GRM7 variants confer susceptibility to age-related hearing impairment

Age-related hearing impairment (ARHI), or presbycusis, is the most prevalent sensory impairment in the elderly. ARHI is a complex disease caused by an interaction between environmental and genetic factors. Here we describe the results of the first whole genome association study for ARHI. The study was performed using 846 cases and 846 controls selected from 3434 individuals collected by eight centers in six European countries. DNA pools for cases and controls were allelotyped on the Affymetrix 500K GeneChip for each center separately. The 252 top-ranked single nucleotide polymorphisms (SNPs) identified in a non-Finnish European sample group (1332 samples) and the 177 top-ranked SNPs from a Finnish sample group (360 samples) were confirmed using individual genotyping. Subsequently, the 23 most interesting SNPs were individually genotyped in an independent European replication group (138 samples). This resulted in the identification of a highly significant and replicated SNP located in GRM7, the gene encoding metabotropic glutamate receptor type 7. Also in the Finnish sample group, two GRM7 SNPs were significant, albeit in a different region of the gene. As the Finnish are genetically distinct from the rest of the European population, this may be due to allelic heterogeneity. We performed histochemical studies in human and mouse and showed that mGluR7 is expressed in hair cells and in spiral ganglion cells of the inner ear. Together these data indicate that common alleles of GRM7 contribute to an individuals risk of developing ARHI, possibly through a mechanism of altered susceptibility to glutamate excitotoxicity.

Genome-wide association analysis demonstrates the highly polygenic character of age-related hearing impairment

We performed a genome-wide association study (GWAS) to identify the genes responsible for age-related hearing impairment (ARHI), the most common form of hearing impairment in the elderly. Analysis of common variants, with and without adjustment for stratification and environmental covariates, rare variants and interactions, as well as gene-set enrichment analysis, showed no variants with genome-wide significance. No evidence for replication of any previously reported genes was found. A study of the genetic architecture indicates for the first time that ARHI is highly polygenic in nature, with probably no major genes involved. The phenotype depends on the aggregated effect of a large number of SNPs, of which the individual effects are undetectable in a modestly powered GWAS. We estimated that 22% of the variance in our data set can be explained by the collective effect of all genotyped SNPs. A score analysis showed a modest enrichment in causative SNPs among the SNPs with a P-value below 0.01.

Genetic evidence for an ethnic diversity in the susceptibility to Ménière's disease.

Background Ménière’s disease (MD) is a debilitating disorder of the inner ear characterized by cochlear and vestibular dysfunction. The cause of this disease is still unknown, and epidemiological data for MD are sparse. From the existing literature, women seem to be more susceptible than men, and Caucasians seem to be more susceptible than Asians. Objective In this article, we characterize a large definite MD cohort for sex and age of onset of disease and use molecular genetic methodologies to characterize ethnicity. Study Design Medical record review for sex and age of onset. Ancestry analysis compared results from the principal component analysis of whole-genome genotype data from MD patients to self-identified ancestry in control samples. Setting House Clinic in Los Angeles. Patients Definitive MD patients. Results Our review of medical records for definitive MD patients reveals that women are more susceptible than men. We also find that men and women have nearly identical age of onset for disease. Lastly, interrogation of molecular genetic data with principal component analysis allowed detailed observations about the ethnic ancestry of our patients. Comparison of the ethnicity of MD patients presenting to our tertiary care clinic with the self-recollected ethnicity of all patients visiting the clinic revealed an ethnic bias, with Caucasians presenting at a higher frequency than expected and the remaining major ethnicities populating Los Angeles (Hispanics, Blacks, and Asians) presenting at a lower frequency than expected. Conclusion To the best of our knowledge, this report is the first ethnic characterization of a large MD cohort from a large metropolitan region using molecular genetic data. Our data suggest that there is a bias in sex and ethnic susceptibility to this disease.

A comprehensive catalogue of the coding and non-coding transcripts of the human inner ear

The mammalian inner ear consists of the cochlea and the vestibular labyrinth (utricle, saccule, and semicircular canals), which participate in both hearing and balance. Proper development and life-long function of these structures involves a highly complex coordinated system of spatial and temporal gene expression. The characterization of the inner ear transcriptome is likely important for the functional study of auditory and vestibular components, yet, primarily due to tissue unavailability, detailed expression catalogues of the human inner ear remain largely incomplete. We report here, for the first time, comprehensive transcriptome characterization of the adult human cochlea, ampulla, saccule and utricle of the vestibule obtained from patients without hearing abnormalities. Using RNA-Seq, we measured the expression of >50,000 predicted genes corresponding to approximately 200,000 transcripts, in the adult inner ear and compared it to 32 other human tissues. First, we identified genes preferentially expressed in the inner ear, and unique either to the vestibule or cochlea. Next, we examined expression levels of specific groups of potentially interesting RNAs, such as genes implicated in hearing loss, long non-coding RNAs, pseudogenes and transcripts subject to nonsense mediated decay (NMD). We uncover the spatial specificity of expression of these RNAs in the hearing/balance system, and reveal evidence of tissue specific NMD. Lastly, we investigated the non-syndromic deafness loci to which no gene has been mapped, and narrow the list of potential candidates for each locus. These data represent the first high-resolution transcriptome catalogue of the adult human inner ear. A comprehensive identification of coding and non-coding RNAs in the inner ear will enable pathways of auditory and vestibular function to be further defined in the study of hearing and balance. Expression data are freely accessible at https://www.tgen.org/home/research/research-divisions/neurogenomics/supplementary-data/inner-ear-transcriptome.aspx.

The Genetic Architecture of Hearing Impairment in Mice: Evidence for Frequency-Specific Genetic Determinants.

Genome-wide association studies (GWAS) have been successfully applied in humans for the study of many complex phenotypes. However, identification of the genetic determinants of hearing in adults has been hampered, in part, by the relative inability to control for environmental factors that might affect hearing throughout the lifetime, as well as a large degree of phenotypic heterogeneity. These and other factors have limited the number of large-scale studies performed in humans that have identified candidate genes that contribute to the etiology of this complex trait. To address these limitations, we performed a GWAS analysis using a set of inbred mouse strains from the Hybrid Mouse Diversity Panel. Among 99 strains characterized, we observed approximately two-fold to five-fold variation in hearing at six different frequencies, which are differentiated biologically from each other by the location in the cochlea where each frequency is registered. Among all frequencies tested, we identified a total of nine significant loci, several of which contained promising candidate genes for follow-up study. Taken together, our results indicate the existence of both genes that affect global cochlear function, as well as anatomical- and frequency-specific genes, and further demonstrate the complex nature of mammalian hearing variation.

Large-scale phenotyping of noise-induced hearing loss in 100 strains of mice.

A cornerstone technique in the study of hearing is the Auditory Brainstem Response (ABR), an electrophysiologic technique that can be used as a quantitative measure of hearing function. Previous studies have published databases of baseline ABR thresholds for mouse strains, providing a valuable resource for the study of baseline hearing function and genetic mapping of hearing traits in mice. In this study, we further expand upon the existing literature by characterizing the baseline ABR characteristics of 100 inbred mouse strains, 47 of which are newly characterized for hearing function. We identify several distinct patterns of baseline hearing deficits and provide potential avenues for further investigation. Additionally, we characterize the sensitivity of the same 100 strains to noise exposure using permanent thresholds shifts, identifying several distinct patterns of noise-sensitivity. The resulting data provides a new resource for studying hearing loss and noise-sensitivity in mice.

FVB/NJ mice demonstrate a youthful sensitivity to noise-induced hearing loss and provide a useful genetic model for the study of neural hearing loss.

The hybrid mouse diversity panel (HMDP), a panel of 100 strains, has been employed in genome wide association studies (GWAS) to study complex traits in mice. Hearing is a complex trait and the CBA/CaJ mouse strain is a widely used model for age-related hearing impairment (ARHI) and noise-induced hearing loss (NIHL). The youthful sensitivity to noise and limited age-related hearing loss of the CBA/CaJ strain led us to attempt the identification of additional strains segregating a similar phenotype for our panel. FVB/NJ is part of the HMDP and has been previously described as having a similar ARHI phenotype to CBA/CaJ. For these reasons, we have studied the FVB/NJ mouse for ARHI and NIHL phenotypes in the hopes of incorporating its phenotype into HMDP studies. We demonstrate that FVB/NJ exhibits ARHI at an earlier age than CBA/CaJ and young FVB/NJ mice are vulnerable to NIHL up to 10-12 weeks. This suggests that FVB/NJ may be used as an additional genetic model for neural forms of progressive hearing loss and for the study of youthful sensitivity to noise.