Ayala Lagziel

Tricellulin Is a Tight-Junction Protein Necessary for Hearing

The inner ear has fluid-filled compartments of different ionic compositions, including the endolymphatic and perilymphatic spaces of the organ of Corti; the separation from one another by epithelial barriers is required for normal hearing. TRIC encodes tricellulin, a recently discovered tight-junction (TJ) protein that contributes to the structure and function of tricellular contacts of neighboring cells in many epithelial tissues. We show that, in humans, four different recessive mutations of TRIC cause nonsyndromic deafness (DFNB49), a surprisingly limited phenotype, given the widespread tissue distribution of tricellulin in epithelial cells. In the inner ear, tricellulin is concentrated at the tricellular TJs in cochlear and vestibular epithelia, including the structurally complex and extensive junctions between supporting and hair cells. We also demonstrate that there are multiple alternatively spliced isoforms of TRIC in various tissues and that mutations of TRIC associated with hearing loss remove all or most of a conserved region in the cytosolic domain that binds to the cytosolic scaffolding protein ZO-1. A wild-type isoform of tricellulin, which lacks this conserved region, is unaffected by the mutant alleles and is hypothesized to be sufficient for structural and functional integrity of epithelial barriers outside the inner ear.

The Tip-Link Antigen, a Protein Associated with the Transduction Complex of Sensory Hair Cells, Is Protocadherin-15

Sound and acceleration are detected by hair bundles, mechanosensory structures located at the apical pole of hair cells in the inner ear. The different elements of the hair bundle, the stereocilia and a kinocilium, are interconnected by a variety of link types. One of these links, the tip link, connects the top of a shorter stereocilium with the lateral membrane of an adjacent taller stereocilium and may gate the mechanotransducer channel of the hair cell. Mass spectrometric and Western blot analyses identify the tip-link antigen, a hitherto unidentified antigen specifically associated with the tip and kinocilial links of sensory hair bundles in the inner ear and the ciliary calyx of photoreceptors in the eye, as an avian ortholog of human protocadherin-15, a product of the gene for the deaf/blindness Usher syndrome type 1F/DFNB23 locus. Multiple protocadherin-15 transcripts are shown to be expressed in the mouse inner ear, and these define four major isoform classes, two with entirely novel, previously unidentified cytoplasmic domains. Antibodies to the three cytoplasmic domain-containing isoform classes reveal that each has a different spatiotemporal expression pattern in the developing and mature inner ear. Two isoforms are distributed in a manner compatible for association with the tip-link complex. An isoform located at the tips of stereocilia is sensitive to calcium chelation and proteolysis with subtilisin and reappears at the tips of stereocilia as transduction recovers after the removal of calcium chelators. Protocadherin-15 is therefore associated with the tip-link complex and may be an integral component of this structure and/or required for its formation.

Recent Advances in the Understanding of Syndromic Forms of Hearing Loss

There are hundreds of different syndromes that include an auditory phenotype as a prominent feature and nearly as many reviews of this topic (Ahmed, Riazuddin, Riazuddin, & Wilcox, 2003; Griffith & Friedman, 2002; Gurtler & Lalwani, 2002; Lalwani, 2002; Morton, 2002; Petit, 2001; Steel, Erven, & Kiernan, 2002). Approximately 30% of individuals with hereditary hearing loss also have abnormalities of other organ systems and are considered to have a syndromic form of deafness (Gorlin, Toriello, & Cohen, 1995). The accompanying abnormalities range from subtle to obvious and may be congenital or delayed in appearance. The majority of these syndromes are inherited as monogenic disorders. Some of these genes have been mapped to a chromosomal map position and a subset of these mapped genes have been identified (cloned). In Table 1 we list many of the syndromic forms of hearing loss and some of the distinguishing clinical features. In many cases where hearing loss is listed as a part of a syndrome, the loss develops late and may simply be secondary to the general neurological decay. For virtually all inherited syndromic forms of hearing loss the Online Mendelian Inheritance in Man (www.ncbi.nlm.nih.gov/Omim/) has comprehensive descriptions of the clinical features and molecular genetics as well as an all-inclusive list of references. In this review we discuss only advances in our understanding of syndromic forms of deafness that have been made in the past few years. Several deafness syndromes are named after the clinician(s) who first or more fully described the disorder such as Jervell and Lange-Nielsen syndrome, Marshall syndrome, Stickler syndrome, Usher syndrome and Waardenburg syndrome (Table 1). Other hereditary deafness syndromes have been given labels that encompass part or all of the clinical presentation (phenotype) such as Branchio-Oto-Renal syndrome (abbreviated BOR, Table 1). As might be expected, there is a gene for each of these clinically distinct inherited syndromes that include hearing loss as one feature. However, there are exceptions to this generalization. Two phenotypically distinct syndromes may be due to different mutations of the same gene. Geneticists refer to different mutations of the same gene as allelic mutations or, more simply, as alleles. There are many examples of phenotypically distinct syndromes that are caused by allelic mutations such as Marshall syndrome (OMIM 154780) and Stickler syndrome type 2 (OMIM 604841), both of which can be caused by mutations of COL11A1 on chromosome 1p21 (Griffith, Sprunger, Sirko-Osadsa, Tiller, Meisler, & Warman, 1998) (Table 1). Another example is Waardenburg syndrome type 1 (OMIM 193500), Waardenburg syndrome type 3 (OMIM 148820) and Craniofacial-Deafness-Hand syndrome (OMIM 122880), which are clinically distinct but, in fact, are caused by allelic mutations of PAX3 on chromosome 2q35 (Asher, Sommer, Morell, & Friedman, 1996). Moreover, the converse is also true. Mutations of more than one gene may cause the identical clinical phenotype. This is referred to as genetic heterogeneity. For example, Usher syndrome type 1 is characterized by congenital, severe to profound hearing loss, retinitis pigmentosa (RP) with prepubertal onset and vestibular areflexia. Surprisingly, there are at least seven different genes that can cause clinically indistinguishable Usher syndrome type 1 (Table 1). Sometimes a patient has hearing loss that is obvious while their other associated abnormalities escape notice. There are many different reasons for incomplete diagnoses. Two examples illustrate this point. The hearing loss in Usher syndrome types 1 and 2 is congenital, while the onset of RP may be delayed and not noticed until adolescence. Patients who have Jervell and Lange-Nielsen syndrome are hearing impaired, but their heart conduction problems are easily overlooked (Table 1). Therefore, Section on Human Genetics (T.B.F., J.M.S., T.B-Y., A.L., E.R.W., S.R., Z.A., I.A.B.), Section on Gene Structure and Function (S.P.P, A.J.G.), and Hearing Section (S.P.P., A.J.G.), National Institute on Deafness and Other Communication Disorders, National Institutes of Health, Rockville, Maryland; and Department of Pediatrics and Human Development (R.A.F.), Michigan State University, East Lansing, Michigan.

A null mutation of mouse Kcna10 causes significant vestibular and mild hearing dysfunction.

KCNA10 is a voltage gated potassium channel that is expressed in the inner ear. The localization and function of KCNA10 was studied in a mutant mouse, B6-Kcna10(TM45), in which the single protein coding exon of Kcna10 was replaced with a beta-galactosidase reporter cassette. Under the regulatory control of the endogenous Kcna10 promoter and enhancers, beta-galactosidase was expressed in hair cells of the vestibular organs and the organ of Corti. KCNA10 expression develops in opposite tonotopic gradients in the inner and outer hair cells. Kcna10(TM45) homozygotes display only a mild elevation in pure tone hearing thresholds as measured by auditory brainstem response (ABR), while heterozygotes are normal. However, Kcna10(TM45) homozygotes have absent vestibular evoked potentials (VsEPs) or elevated VsEP thresholds with prolonged peak latencies, indicating significant vestibular dysfunction despite the lack of any overt imbalance behaviors. Our results suggest that Kcna10 is expressed primarily in hair cells of the inner ear, with little evidence of expression in other organs. The Kcna10(TM45) targeted allele may be a model of human nonsyndromic vestibulopathy.

Modifier variant of METTL13 suppresses human GAB1–associated profound deafness

A modifier variant can abrogate the risk of a monogenic disorder. DFNM1 is a locus on chromosome 1 encoding a dominant suppressor of human DFNB26 recessive, profound deafness. Here, we report that DFNB26 is associated with a substitution (p.Gly116Glu) in the pleckstrin homology domain of GRB2-associated binding protein 1 (GAB1), an essential scaffold in the MET proto-oncogene, receptor tyrosine kinase/HGF (MET/HGF) pathway. A dominant substitution (p.Arg544Gln) of METTL13, encoding a predicted methyltransferase, is the DFNM1 suppressor of GAB1-associated deafness. In zebrafish, human METTL13 mRNA harboring the modifier allele rescued the GAB1-associated morphant phenotype. In mice, GAB1 and METTL13 colocalized in auditory sensory neurons, and METTL13 coimmunoprecipitated with GAB1 and SPRY2, indicating at least a tripartite complex. Expression of MET-signaling genes in human lymphoblastoid cells of individuals homozygous for p.Gly116Glu GAB1 revealed dysregulation of HGF, MET, SHP2, and SPRY2, all of which have reported variants associated with deafness. However, SPRY2 was not dysregulated in normal-hearing humans homozygous for both the GAB1 DFNB26 deafness variant and the dominant METTL13 deafness suppressor, indicating a plausible mechanism of suppression. Identification of METTL13-based modification of MET signaling offers a potential therapeutic strategy for a wide range of associated hearing disorders. Furthermore, MET signaling is essential for diverse functions in many tissues including the inner ear. Therefore, identification of the modifier of MET signaling is likely to have broad clinical implications.