9.A.46 The Clarin (CLRN) Family
Usher syndrome type 3 (USH3) is an autosomal recessive disorder characterized by the association of post-lingual progressive hearing loss, progressive visual loss due to retinitis pigmentosa and variable presence of vestibular dysfunction (Adato et al., 2002). The full length USH3A transcript encodes clarin-1 (CLRN1), a four-transmembrane-domain protein of the tetraspanin family, with 3 vertebrate paralogues, Clarin-1, 2 and 3. Limited sequence homology to stargazin, a cerebellar synapse four-transmembrane-domain protein, suggested a role for clarin-1 in hair cell and photoreceptor cell synapses, as well as a common pathophysiological pathway for different Usher syndromes (Adato et al., 2002). CLRN1 is nonessential in the mouse retina but is required for cochlear hair cell development (Geller et al., 2009). Cones were observed centrally but not in regions with scotomas, and retinal pigment epithelial cells were visible in regions without cones in patients with CLRN1 mutations.
Mice carrying null mutations in the clarin-1 gene (Clrn1(-/-)) show loss of hair cell function. Immunohistochemistry and patch-clamp recordings of Ca2 currents and membrane capacitance from inner hair cells showed that clarin-1 is not essential for formation or function of ribbon synapses. However, reduced cochlear microphonic potentials, FM1-43 [N-(3-triethylammoniumpropyl)-4-(4-(dibutylamino)styryl) pyridinium dibromide] loading, and transduction currents pointed to diminished cochlear hair bundle function in Clrn1(-/-) mice. Electron microscopy of cochlear hair cells revealed loss of some tall stereocilia and gaps in the v-shaped bundle, although tip links and staircase arrangement of stereocilia were not primarily affected by Clrn1(-/-) mutations. Human clarin-1 protein expressed in transfected mouse cochlear hair cells localized to the bundle; however, the pathogenic variant p.N48K failed to localize to the bundle. Thus, CLRN1 is an essential hair bundle protein (Geller et al. 2009).
The molecular mechanisms underlying hair cell synaptic maturation are not well understood. Cadherin-23 (CDH23), protocadherin-15 (PCDH15) and the very large G-protein coupled receptor 1 (VLGR1) have been implicated in the development of cochlear hair cell stereocilia. Mutations in CDH23, PCDH15, VLGR1 and clarin-1 cause Usher syndrome, characterized by congenital deafness, vestibular dysfunction and retinitis pigmentosa. Zallocchi et al. (2012) demonstrated developmental expression of these Usher proteins in afferent spiral ganglion neurons and hair cell synapses. They identified a novel synaptic Usher complex comprised of clarin-1 and specific isoforms of CDH23, PCDH15 and VLGR1. To establish the in vivo relevance of this complex, they performed morphological and quantitative analyses of the neuronal fibers and their synapses in the Clrn1-/- mouse. Thus, usher protein complexes comprised of specific protein isoforms probably function in synaptic maturation.