A Novel C-Terminal CIB2 (Calcium and Integrin Binding Protein 2) Mutation Associated with Non-Syndromic Hearing Loss in a Hispanic Family

Hearing loss is a complex disorder caused by both genetic and environmental factors. Previously, mutations in CIB2 have been identified as a common cause of genetic hearing loss in Pakistani and Turkish populations. Here we report a novel (c.556C\u3eT; p.(Arg186Trp)) transition mutation in the CIB2 gene identified through whole exome sequencing (WES) in a Caribbean Hispanic family with non-syndromic hearing loss. CIB2 belongs to the family of calcium-and integrin-binding (CIB) proteins. The carboxy-termini of CIB proteins are associated with calcium binding and intracellular signaling. The p.(Arg186Trp) mutation is localized within predicted type II PDZ binding ligand at the carboxy terminus. Our ex vivo studies revealed that the mutation did not alter the interactions of CIB2 with Whirlin, nor its targeting to the tips of hair cell stereocilia. However, we found that the mutation disrupts inhibition of ATP-induced Ca2+ responses by CIB2 in a heterologous expression system. Our findings support p.(Arg186Trp) mutation as a cause for hearing loss in this Hispanic family. In addition, it further highlights the necessity of the calcium binding property of CIB2 for normal hearing

CIB2 Interacts with TMC1 and TMC2 and is Essential for Mechanotransduction in Auditory Hair Cells

Inner ear hair cells detect sound through deflection of stereocilia, the microvilli-like projections that are arranged in rows of graded heights. Calcium and integrin-binding protein 2 is essential for hearing and localizes to stereocilia, but its exact function is unknown. Here, we have characterized two mutant mouse lines, one lacking calcium and integrin-binding protein 2 and one carrying a human deafness-related Cib2 mutation, and show that both are deaf and exhibit no mechanotransduction in auditory hair cells, despite the presence of tip links that gate the mechanotransducer channels. In addition, mechanotransducing shorter row stereocilia overgrow in hair cell bundles of both Cib2 mutants. Furthermore, we report that calcium and integrin-binding protein 2 binds to the components of the hair cell mechanotransduction complex, TMC1 and TMC2, and these interactions are disrupted by deafness-causing Cib2 mutations. We conclude that calcium and integrin-binding protein 2 is required for normal operation of the mechanotransducer channels and is involved in limiting the growth of transducing stereocilia

PLoS Pathog

Cytomegalovirus (CMV) is a leading infectious cause of morbidity in immune-compromised patients. γδ T cells have been involved in the response to CMV but their role in protection has not been firmly established and their dependency on other lymphocytes has not been addressed. Using C57BL/6 αβ and/or γδ T cell-deficient mice, we here show that γδ T cells are as competent as αβ T cells to protect mice from CMV-induced death. γδ T cell-mediated protection involved control of viral load and prevented organ damage. γδ T cell recovery by bone marrow transplant or adoptive transfer experiments rescued CD3ε-/- mice from CMV-induced death confirming the protective antiviral role of γδ T cells. As observed in humans, different γδ T cell subsets were induced upon CMV challenge, which differentiated into effector memory cells. This response was observed in the liver and lungs and implicated both CD27+ and CD27- γδ T cells. NK cells were the largely preponderant producers of IFNγ and cytotoxic granules throughout the infection, suggesting that the protective role of γδ T cells did not principally rely on either of these two functions. Finally, γδ T cells were strikingly sufficient to fully protect Rag-/-γc-/- mice from death, demonstrating that they can act in the absence of B and NK cells. Altogether our results uncover an autonomous protective antiviral function of γδ T cells, and open new perspectives for the characterization of a non classical mode of action which should foster the design of new γδ T cell based therapies, especially useful in αβ T cell compromised patients

Radioprotective Effect of Aminothiol PrC-210 on Irradiated Inner Ear of Guinea Pig.

Radiotherapy of individuals suffering with head & neck or brain tumors subserve the risk of sensorineural hearing loss. Here, we evaluated the protective effect of Aminothiol PrC-210 (3-(methyl-amino)-2-((methylamino)methyl)propane-1-thiol) on the irradiated inner ear of guinea pigs. An intra-peritoneal or intra-tympanic dose of PrC-210 was administered prior to receiving a dose of gamma radiation (3000 cGy) to each ear. Auditory Brainstem Responses (ABRs) were recorded one week and two weeks after the radiation and compared with the sham animal group. ABR thresholds of guinea pigs that received an intra-peritoneal dose of PrC-210 were significantly better compared to the non-treated, control animals at one week post-radiation. Morphologic analysis of the inner ear revealed significant inflammation and degeneration of the spiral ganglion in the irradiated animals not treated with PrC-210. In contrast, when treated with PrC-210 the radiation effect and injury to the spiral ganglion was significantly alleviated. PrC-210 had no apparent cytotoxic effect in vivo and did not affect the morphology or count of cochlear hair cells. These findings suggest that aminothiol PrC-210 attenuated radiation-induced cochlea damage for at least one week and protected hearing

Auditory brainstem response (ABR) thresholds of irradiated adult guinea pigs treated by intra-peritoneal or intra-tympanic injection of PrC-210.

<p>(A) Total average thresholds at day 0, week 1 and week 2. Radiation: animals received 3000 cGy in each ear. Non-treated animals (red bars), animals treated with intra-peritoneal injection of PrC-210 (blue bars), animals treated with intra-tympanic injection of PrC-210 (yellow bars). (B) Hearing thresholds shift for the left and right ears of irradiated adult guinea pigs not treated with PrC-210 (Red) and treated by intra-peritoneal injection of PrC-210 (Blue). Total average hearing loss in dB at week 1 (light blue and light red bars) and week 2 (dark blue and dark red bars). (C) ABR pure tone thresholds of irradiated adult guinea pigs treated by an intra-peritoneal or intra-tympanic injection of PrC-210. Average pure tone thresholds at day 0, week 1 and week 2 measured at 8kHz, 16kHz and 32kHz. Radiation (Rad): animals received 3000 cGy in each ear. IP: intra-peritoneal injection of PrC-210 (blue triangles), IT: intra-tympanic injection of PrC-210 (inverted yellow triangles). Non-treated animals are indicated with red squares. Decibels (dB), *, p< = 0.05; **, p< = 0.01; ***, p< = 0.001, n.s., non significant.</p

The aminothiol PrC-210 prevents the inflammation and the degeneration of the apical part of the spiral ganglion induced by the radiation.

<p>Hematoxylin and Eosin staining of paraffin section of guinea pig cochleae untreated and treated by intra-peritoneal injection of PrC-210 before receiving a 3000 cGy radiation dose. Guinea pig cochleae from radiation only and PrC-210 IP groups were harvest at two weeks after being irradiated. Arrowheads indicate the sensory hair cells in the organ of Corti. Scale bar: 100μm.</p

Cochlear sensory hair cell death in irradiated adult guinea pigs treated by an intra-peritoneal injection of PrC-210.

<p>(A) Cochleae from irradiated adult guinea pigs that were treated by an intra-peritoneal (IP) injection of PrC-210 were dissected at week 2. Cochleae were immunostained with Myosin VIIa antibody (Green), a hair cell marker and phalloidin (Red, Grey scale), an actin marker. Scale bar = 100 μm. Dead hair cells are labeled with a star (*) (B) The dead sensory hair cells were quantified and compared to the control group. n.s., non significant.</p

Radiation shielding devices.

<p>Diagram showing the lead shield that was used in order to protect the sedated guinea pigs. A 4x¾ “opening was drilled, so that the ear/auricle region of the guinea was exposed to radiation. Guinea pigs were irradiated with a Cs-137 irradiator. Sedated guinea pigs were placed on an elevated stage at the second position of the irradiator, behind the lead shield.</p

Distribution of Guinea pigs into various study groups.

<p>Distribution of Guinea pigs into various study groups.</p

Gipc1 has a dual role in Vangl2 trafficking and hair bundle integrity in the inner ear

International audienceVangl2 is one of the central proteins controlling the establishment of planar cell polarity in multiple tissues of different species. Previous studies suggest that the localization of the Vangl2 protein to specific intracellular microdomains is crucial for its function. However, the molecular mechanisms that control Vangl2 trafficking within a cell are largely unknown. Here, we identify Gipc1 (GAIP C-terminus interacting protein 1) as a new interactor for Vangl2, and we show that a myosin VI-Gipc1 protein complex can regulate Vangl2 traffic in heterologous cells. Furthermore, we show that in the cochlea of MyoVI mutant mice, Vangl2 presence at the membrane is increased, and that a disruption of Gipc1 function in hair cells leads to maturation defects, including defects in hair bundle orientation and integrity. Finally, stimulated emission depletion microscopy and overexpression of GFP-Vangl2 show an enrichment of Vangl2 on the supporting cell side, adjacent to the proximal membrane of hair cells. Altogether, these results indicate a broad role for Gipc1 in the development of both stereociliary bundles and cell polarization, and suggest that the strong asymmetry of Vangl2 observed in early postnatal cochlear epithelium is mostly a ‘tissue’ polarity readout