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1.  The CD2 isoform of protocadherin-15 is an essential component of the tip-link complex in mature auditory hair cells 
EMBO Molecular Medicine  2014;6(7):984-992.
Protocadherin-15 (Pcdh15) is a component of the tip-links, the extracellular filaments that gate hair cell mechano-electrical transduction channels in the inner ear. There are three Pcdh15 splice isoforms (CD1, CD2 and CD3), which only differ by their cytoplasmic domains; they are thought to function redundantly in mechano-electrical transduction during hair-bundle development, but whether any of these isoforms composes the tip-link in mature hair cells remains unknown. By immunolabelling and both morphological and electrophysiological analyses of post-natal hair cell-specific conditional knockout mice (Pcdh15ex38-fl/ex38-fl Myo15-cre+/−) that lose only this isoform after normal hair-bundle development, we show that Pcdh15-CD2 is an essential component of tip-links in mature auditory hair cells. The finding, in the homozygous or compound heterozygous state, of a PCDH15 frameshift mutation (p.P1515Tfs*4) that affects only Pcdh15-CD2, in profoundly deaf children from two unrelated families, extends this conclusion to humans. These results provide key information for identification of new components of the mature auditory mechano-electrical transduction machinery. This will also serve as a basis for the development of gene therapy for deafness caused by PCDH15 defects.
PMCID: PMC4119359  PMID: 24940003
auditory mechano-electrical transduction; deafness; protocadherin-15; stereocilia; tip-link
2.  Targeted Ablation of Connexin26 in the Inner Ear Epithelial Gap Junction Network Causes Hearing Impairment and Cell Death 
Current biology : CB  2002;12(13):1106-1111.
Mutations in the gene encoding the gap junction protein connexin26 (Cx26) are responsible for the autosomal recessive isolated deafness, DFNB1, which accounts for half of the cases of prelingual profound hereditary deafness in Caucasian populations [1–5]. To date, in vivo approaches to decipher the role of Cx26 in the inner ear have been hampered by the embryonic lethality of the Cx26 knockout mice [6]. To overcome this difficulty, we performed targeted ablation of Cx26 specifically in one of the two cellular networks that it underlies in the inner ear, namely, the epithelial network. We show that homozygous mutant mice, Cx26OtogCre, have hearing impairment, but no vestibular dysfunction. The inner ear developed normally. However, on postnatal day 14 (P14), i.e., soon after the onset of hearing, cell death appeared and eventually extended to the cochlear epithelial network and sensory hair cells. Cell death initially affected only the supporting cells of the genuine sensory cell (inner hair cell, IHC), thus suggesting that it could be triggered by the IHC response to sound stimulation. Altogether, our results demonstrate that the Cx26-containing epithelial gap junction network is essential for cochlear function and cell survival. We conclude that prevention of cell death in the sensory epithelium is essential for any attempt to restore the auditory function in DFNB1 patients.
PMCID: PMC4030438  PMID: 12121617
3.  Localization of Usher 1 proteins to the photoreceptor calyceal processes, which are absent from mice 
The Journal of Cell Biology  2012;199(2):381-399.
Mice are a poor model for retinal defects caused by type I Usher syndrome (USH1) because their photoreceptors have almost no calyceal processes, the structures in which all USH1 proteins are detected in other vertebrates.
The mechanisms underlying retinal dystrophy in Usher syndrome type I (USH1) remain unknown because mutant mice lacking any of the USH1 proteins—myosin VIIa, harmonin, cadherin-23, protocadherin-15, sans—do not display retinal degeneration. We found here that, in macaque photoreceptor cells, all USH1 proteins colocalized at membrane interfaces (i) between the inner and outer segments in rods and (ii) between the microvillus-like calyceal processes and the outer segment basolateral region in rods and cones. This pattern, conserved in humans and frogs, was mediated by the formation of an USH1 protein network, which was associated with the calyceal processes from the early embryonic stages of outer segment growth onwards. By contrast, mouse photoreceptors lacked calyceal processes and had no USH1 proteins at the inner–outer segment interface. We suggest that USH1 proteins form an adhesion belt around the basolateral region of the photoreceptor outer segment in humans, and that defects in this structure cause the retinal degeneration in USH1 patients.
PMCID: PMC3471240  PMID: 23045546
4.  Coupling of the mechanotransduction machinery and F-actin polymerization in the cochlear hair bundles 
Bioarchitecture  2011;1(4):169-174.
Mechanoelectrical transduction (MET), the conversion of mechanical stimuli into electrical signals operated by the sensory cells of the inner ear, enables hearing and balance perception. Crucial to this process are the tip-links, oblique fibrous filaments that interconnect the actin-filled stereocilia of different rows within the hair bundle, and mechanically gate MET channels. In a recent study, we observed a complete regression of stereocilia from the short and medium but not the tall row upon the disappearance of the tip-links caused by the loss of one of their components, cadherin-23, or of one of their anchoring proteins, sans, in the auditory organs of engineered mutant mice. This indicates the existence of a coupling between the MET and F-actin polymerization machineries at the tips of the short and medium stereocilia rows in cochlear hair bundles. Here, we first present our findings in the mutant mice, and then discuss the possible effects of the tip-link tension on stereocilia F-actin polymerization, acting either directly or through Ca2+-dependent mechanisms that involve the gating of MET channels.
PMCID: PMC3210516  PMID: 22069509
hair cell; hair bundle; stereocilia; mechanoelectrical transduction (MET); tip-link; sans protein; actin polymerization
5.  Total variation regularization for fMRI-based prediction of behavior 
Ieee Transactions on Medical Imaging  2011;30(7):1328-1340.
While medical imaging typically provides massive amounts of data, the extraction of relevant information for predictive diagnosis remains a difficult challenge. Functional MRI (fMRI) data, that provide an indirect measure of task-related or spontaneous neuronal activity, are classically analyzed in a mass-univariate procedure yielding statistical parametric maps. This analysis framework disregards some important principles of brain organization: population coding, distributed and overlapping representations. Multivariate pattern analysis, i.e., the prediction of behavioural variables from brain activation patterns better captures this structure. To cope with the high dimensionality of the data, the learning method has to be regularized. However, the spatial structure of the image is not taken into account in standard regularization methods, so that the extracted features are often hard to interpret. More informative and interpretable results can be obtained with the ℓ1 norm of the image gradient, a.k.a. its Total Variation (TV), as regularization. We apply for the first time this method to fMRI data, and show that TV regularization is well suited to the purpose of brain mapping while being a powerful tool for brain decoding. Moreover, this article presents the first use of TV regularization for classification.
PMCID: PMC3336110  PMID: 21317080
fMRI; regression; classification; regularization; Total Variation; spatial structure; Algorithms; Behavior; physiology; Brain; anatomy & histology; physiology; Brain Mapping; Cognition; physiology; Computer Simulation; Humans; Image Processing, Computer-Assisted; methods; Magnetic Resonance Imaging; methods; Multivariate Analysis; Regression Analysis
6.  Multiclass Sparse Bayesian Regression for fMRI-Based Prediction 
Inverse inference has recently become a popular approach for analyzing neuroimaging data, by quantifying the amount of information contained in brain images on perceptual, cognitive, and behavioral parameters. As it outlines brain regions that convey information for an accurate prediction of the parameter of interest, it allows to understand how the corresponding information is encoded in the brain. However, it relies on a prediction function that is plagued by the curse of dimensionality, as there are far more features (voxels) than samples (images), and dimension reduction is thus a mandatory step. We introduce in this paper a new model, called Multiclass Sparse Bayesian Regression (MCBR), that, unlike classical alternatives, automatically adapts the amount of regularization to the available data. MCBR consists in grouping features into several classes and then regularizing each class differently in order to apply an adaptive and efficient regularization. We detail these framework and validate our algorithm on simulated and real neuroimaging data sets, showing that it performs better than reference methods while yielding interpretable clusters of features.
PMCID: PMC3132985  PMID: 21754916
7.  Cadherin-23, myosin VIIa and harmonin, encoded by Usher syndrome type I genes, form a ternary complex and interact with membrane phospholipids 
Human Molecular Genetics  2010;19(18):3557-3565.
Cadherin-23 is a component of early transient lateral links of the auditory sensory cells' hair bundle, the mechanoreceptive structure to sound. This protein also makes up the upper part of the tip links that control gating of the mechanoelectrical transduction channels. We addressed the issue of the molecular complex that anchors these links to the hair bundle F-actin core. By using surface plasmon resonance assays, we show that the cytoplasmic regions of the two cadherin-23 isoforms that do or do not contain the exon68-encoded peptide directly interact with harmonin, a submembrane PDZ (post-synaptic density, disc large, zonula occludens) domain-containing protein, with unusually high affinity. This interaction involves the harmonin Nter-PDZ1 supramodule, but not the C-terminal PDZ-binding motif of cadherin-23. We establish that cadherin-23 directly binds to the tail of myosin VIIa. Moreover, cadherin-23, harmonin and myosin VIIa can form a ternary complex, which suggests that myosin VIIa applies tension forces on hair bundle links. We also show that the cadherin-23 cytoplasmic region, harmonin and myosin VIIa interact with phospholipids on synthetic liposomes. Harmonin and the cytoplasmic region of cadherin-23, both independently and as a binary complex, can bind specifically to phosphatidylinositol 4,5-bisphosphate (PI(4,5)P2), which may account for the role of this phospholipid in the adaptation of mechanoelectrical transduction in the hair bundle. The distributions of cadherin-23, harmonin, myosin VIIa and PI(4,5)P2 in the growing and mature auditory hair bundles as well as the abnormal locations of harmonin and myosin VIIa in cadherin-23 null mutant mice strongly support the functional relevance of these interactions.
PMCID: PMC2928128  PMID: 20639393
8.  Human adenylate kinase 2 deficiency causes a profound haematopoietic defect associated with sensorineural deafness 
Nature genetics  2008;41(1):106-111.
Reticular dysgenesis (RD) is an autosomal recessive form of human Severe Combined Immunodeficiency, characterized by an early differentiation arrest in the myeloid lineage and impaired lymphoid maturation. In addition, affected newborns have bilateral sensorineural deafness. We have identified biallelic mutations in the adenylate kinase 2 (AK2) gene in seven patients affected with RD. These mutations resulted in the absence or a strong decrease in protein expression. We then demonstrated that restoration of AK2 expression in the bone marrow cells of RD patients overcomes the neutrophil differentiation arrest underlining its specific requirement in the development of a restricted set of haematopoietic lineages. Lastly, we established that AK2 is specifically expressed in the stria vascularis region of the inner ear, which provides an explanation to the sensorineural deafness. These results suggest a novel mechanism regulating haematopoetic cell differentiation, and involved in one of the most severe human immunodeficiency syndromes.
PMCID: PMC2612090  PMID: 19043416
9.  Harmonin-b, an actin-binding scaffold protein, is involved in the adaptation of mechanoelectrical transduction by sensory hair cells 
Pflugers Archiv   2009;459(1):115-130.
We assessed the involvement of harmonin-b, a submembranous protein containing PDZ domains, in the mechanoelectrical transduction machinery of inner ear hair cells. Harmonin-b is located in the region of the upper insertion point of the tip link that joins adjacent stereocilia from different rows and that is believed to gate transducer channel(s) located in the region of the tip link's lower insertion point. In Ush1cdfcr-2J/dfcr-2J mutant mice defective for harmonin-b, step deflections of the hair bundle evoked transduction currents with altered speed and extent of adaptation. In utricular hair cells, hair bundle morphology and maximal transduction currents were similar to those observed in wild-type mice, but adaptation was faster and more complete. Cochlear outer hair cells displayed reduced maximal transduction currents, which may be the consequence of moderate structural anomalies of their hair bundles. Their adaptation was slower and displayed a variable extent. The latter was positively correlated with the magnitude of the maximal transduction current, but the cells that showed the largest currents could be either hyperadaptive or hypoadaptive. To interpret our observations, we used a theoretical description of mechanoelectrical transduction based on the gating spring theory and a motor model of adaptation. Simulations could account for the characteristics of transduction currents in wild-type and mutant hair cells, both vestibular and cochlear. They led us to conclude that harmonin-b operates as an intracellular link that limits adaptation and engages adaptation motors, a dual role consistent with the scaffolding property of the protein and its binding to both actin filaments and the tip link component cadherin-23.
Electronic supplementary material
The online version of this article (doi:10.1007/s00424-009-0711-x) contains supplementary material, which is available to authorized users.
PMCID: PMC2767239  PMID: 19756723
Cochlea; Hair bundle; Vestibule; Harmonin; Mechanoelectrical transduction; Adaptation; Hair cell; Vestibular system
10.  Vezatin, an integral membrane protein of adherens junctions, is required for the sound resilience of cochlear hair cells 
EMBO Molecular Medicine  2009;1(2):125-138.
Loud sound exposure is a significant cause of hearing loss worldwide. We asked whether a lack of vezatin, an ubiquitous adherens junction protein, could result in noise-induced hearing loss. Conditional mutant mice bearing non-functional vezatin alleles only in the sensory cells of the inner ear (hair cells) indeed exhibited irreversible hearing loss after only one minute exposure to a 105 dB broadband sound. In addition, mutant mice spontaneously underwent late onset progressive hearing loss and vestibular dysfunction related to substantial hair cell death. We establish that vezatin is an integral membrane protein with two adjacent transmembrane domains, and cytoplasmic N- and C-terminal regions. Late recruitment of vezatin at junctions between MDCKII cells indicates that the protein does not play a role in the formation of junctions, but rather participates in their stability. Moreover, we show that vezatin directly interacts with radixin in its actin-binding conformation. Accordingly, we provide evidence that vezatin associates with actin filaments at cell–cell junctions. Our results emphasize the overlooked role of the junctions between hair cells and their supporting cells in the auditory epithelium resilience to sound trauma.
PMCID: PMC3378116  PMID: 20049712
adherens junction; mouse model; noise-induced hearing loss; organ of Corti; vezatin

Results 1-10 (10)