Vertebrate hearing and balance are based in complex asymmetries of inner ear structure. Here, we identify retinoic acid (RA) as an extrinsic signal that acts directly on the ear rudiment to affect its compartmentalization along the anterior-posterior axis. A rostrocaudal wave of RA activity, generated by tissues surrounding the nascent ear, induces distinct responses from anterior and posterior halves of the inner ear rudiment. Prolonged response to RA by posterior otic tissue correlates with Tbx1 transcription and formation of mostly nonsensory inner ear structures. By contrast, anterior otic tissue displays only a brief response to RA and forms neuronal elements and most sensory structures of the inner ear.axial specification | developmental compartments | morphogen N ormal hearing and balance require that discrete patches of mechanosensory hair cells, each with a distinct function, be precisely positioned within the asymmetric membranous labyrinth of the inner ear (Fig. 1A). Five vestibular sensory patches are present in all vertebrate inner ears: the three cristae (anterior, lateral, and posterior) that detect angular head movements and two maculae (utricle and saccule) that detect linear acceleration. The specialized organ for detecting sound in chickens and mammals is the basilar papilla and organ of Corti, respectively.The entire membranous labyrinth and its innervating neurons are derived from an ectodermal thickening adjacent to the hindbrain known as the otic placode. As the placode deepens to form a cup and then pinches off to form the otocyst, some cells of the otic epithelium delaminate to form neuroblasts of the cochleovestibular ganglion (CVG). Inner ear sensory organs, and the neurons that innervate them, are thought to arise from a neural-sensory competent domain (NSD), most of which is located in the anterior region of the otic cup (1). By contrast, posterior otic epithelium forms nonsensory tissues and only one sensory organ, the posterior crista. This basic organization of functional elements in the ear is thought to be governed by signals emanating from adjacent tissues (2, 3); however, molecular mechanisms that establish the initial anterior-posterior (A-P) asymmetry of the ear primordium are poorly defined. Here, we show that a rostrocaudal wave of retinoic acid activity provides signals to the ear rudiment and establishes structural asymmetries required for normal hearing and balance. ResultsEctoderm Adjacent to the Otic Cup Confers A-P Polarity to the Otocyst. A clear manifestation of A-P asymmetry in developing amniote ears is the anterior expression of transcripts associated with cochleovestibular ganglion neurogenesis. We performed tissue transplantations in ovo to identify source(s) of signals that specify the otic A-P axis in the chicken. Transplantations were carried out at the otic cup stage (11-15 somite stages), before the otic A-P axis is specified (4). As expected, reversing the A-P orientation of the otic cup alone resulted in a high occurrence of otocysts with the axial plan...

UK. Clinical and molecular characterizations of novel POU3F4 mutations reveal that DFN3 is due to null function of POU3F4 protein. Physiol Genomics 39: 195-201, 2009. First published August 11, 2009 doi:10.1152/physiolgenomics.00100.2009.-X-linked deafness type 3 (DFN3), the most prevalent X-linked form of hereditary deafness, is caused by mutations in the POU3F4 locus, which encodes a member of the POU family of transcription factors. Despite numerous reports on clinical evaluations and genetic analyses describing novel POU3F4 mutations, little is known about how such mutations affect normal functions of the POU3F4 protein and cause inner ear malformations and deafness. Here we describe three novel mutations of the POU3F4 gene and their clinical characterizations in three Korean families carrying deafness segregating at the DFN3 locus. The three mutations cause a substitution (p.Arg329Pro) or a deletion (p.Ser310del) of highly conserved amino acid residues in the POU homeodomain or a truncation that eliminates both DNA-binding domains (p.Ala116fs). In an attempt to better understand the molecular mechanisms underlying their inner ear defects, we examined the behavior of the normal and mutant forms of the POU3F4 protein in C3H/10T1/2 mesodermal cells. Protein modeling as well as in vitro assays demonstrated that these mutations are detrimental to the tertiary structure of the POU3F4 protein and severely affect its ability to bind DNA. All three mutated POU3F4 proteins failed to transactivate expression of a reporter gene. In addition, all three failed to inhibit the transcriptional activity of wild-type proteins when both wildtype and mutant proteins were coexpressed. Since most of the mutations reported for DFN3 thus far are associated with regions that encode the DNA binding domains of POU3F4, our results strongly suggest that the deafness in DFN3 patients is largely due to the null function of POU3F4. hearing loss; X-linked deafness type 3; inner ear CONGENITAL HEARING LOSS IS one of the most common sensory disorders in humans, affecting approximately one in 1,000 newborns (24). More than 50% of the congenital hearing loss is due to genetic causes, and the majority of these hereditary cases are nonsyndromic. While most genetic nonsyndromic hearing loss is caused by mutations in autosomal genes, X-linked cases are estimated to comprise between 1 and 5% of these (26). Thus far, four different X-linked nonsyndromic hearing loss loci (DFN2, DFN3, DFN4, and DFN6) have been mapped, but the causative gene has been identified only for the DFN3 locus (33).X-linked deafness type 3 (DFN3) accounts for ϳ50% of all families carrying X-linked nonsyndromic hearing loss (26). Clinical characteristics of DFN3 in affected males include temporal bone abnormalities, stapes fixation, and, in most cases, a mixed type of hearing loss, which is often progressive (7,8,10). Anatomical anomalies of the temporal bone revealed by computer-assisted tomography (CT) include dilatation of the lateral end of the internal acoustic canal (IAC...

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A novel gene Jpk (Jopock) has been originally isolated through yeast 1 hybridization technique as a trans-acting factor interacting with the position-specific regulatory element of a murine Hoxa-7. Northern analysis revealed that the Jpk was expressed at day 7.0 post coitum (p.c.) during early gastrulation. Previously it has been shown that a trace amount of JPK protein led bacterial cells to death. In eukaryotic F9 cells, Jpk also led the cell to death-generating DNA ladder: fewer than 50% of the cells survived after 72-h transfection. Flow cytometric analysis with cells stained with each Annexin V/7-amino-actinomycin D (7-AAD), MitoTracker, and hydroethidine (HE) revealed that Jpk induced apoptotic cell death in a time-dependent manner, reduced mitochondrial membrane potential, and increased ROS (reactive oxygen species) production, respectively. Additionally, Jpk seemed to regulate the Bcl family at the transcriptional level when RT-PCR was performed. Although the precise mechanism is not clear, these results altogether suggest that Jpk is a potent inducer of apoptosis through generation of ROS as well as concomitant reduction of mitochondrial membrane potential.