Dispensability of Tubulin Acetylation for 15-protofilament Microtubule Formation in the Mammalian Cochlea.

The development of hearing in mammals requires the formation and maturation of a highly organized and specialized epithelium known as the organ of Corti. This epithelium contains two types of cells, the sensory cells, which are the true receptors of auditory information, and the surrounding supporting cells, which are composed of a highly developed cytoskeleton essential to the architecture of the mature organ of Corti. The supporting cells are the only mammalian cells reported to contain the unusual 15-protofilament microtubules. In this paper, we show that 15-protofilament microtubules appear between the second and fourth day after birth in the pillar cells of the organ of Corti in mice. We also show that contrary to what has been described in the nematode worm Caenorhabiditis. elegans, microtubule acetylation is not essential for the formation of 15-protofilament microtubules in mice but is required for fine-tuning of their diameter.Key words: Acetylation, cytoskeleton, microtubule, inner ear, supporting cells.

Relationships between the structural and functional organization of the turtle cell nucleolus.

The nucleolus is a multifunctional structure of the eukaryotic cell nucleus. However, its primary role is ribosome formation. Although the factors and mechanisms involved in ribogenesis are well conserved in eukaryotes, two types of nucleoli have been observed under the electron microscope: a tricompartmentalized nucleolus in amniotes and a bicompartmentalized nucleolus in other species. A recent study has also revealed that turtles, although belonging to amniotes, displayed a nucleolus with bipartite organization, suggesting that this reptile group may have carried out a reversion phenomenon during evolution. In this study, we examine in great detail the functional organization of the turtle nucleolus. In liver and spleen cells cultured in vitro, we confirm that the turtle nucleolus is mainly formed by two components: a fibrillar zone surrounded by a granular zone. We further show that the fibrillar zone includes densely-contrasted strands, which are positive after silver-stained Nucleolar Organizer Region (Ag-NOR) staining and DNA labelling. We also reveal that the dense strands condensed into a very compact mass within the fibrillar zone after a treatment with actinomycin D or 5,6-dichlorobenzimidazole riboside. Finally, by using pulse-chase experiments with BrUTP, three-dimensional image reconstructions of confocal optical sections, and electron microscopy analysis of ultrathin sections, we show that the topological and spatial dynamics of rRNA within the nucleolus extend from upstream binding factor (UBF)-positive sites in the fibrillar zone to the granular zone, without ever releasing the positive sites for the UBF. Together, these results seem to clearly indicate that the compartmentalization of the turtle nucleolus into two main components reflects a less orderly organization of ribosome formation.

MacroH2A histone variants maintain nuclear organization and heterochromatin architecture.

Genetic loss-of-function studies on development, cancer and somatic cell reprogramming have suggested that the group of macroH2A histone variants might function through stabilizing the differentiated state by a yet unknown mechanism. Here, we present results demonstrating that macroH2A variants have a major function in maintaining nuclear organization and heterochromatin architecture. Specifically, we find that a substantial amount of macroH2A is associated with heterochromatic repeat sequences. We further identify macroH2A on sites of interstitial heterochromatin decorated by histone H3 trimethylated on K9 (H3K9me3). Loss of macroH2A leads to major defects in nuclear organization, including reduced nuclear circularity, disruption of nucleoli and a global loss of dense heterochromatin. Domains formed by DNA repeat sequences are disorganized, expanded and fragmented, and mildly re-expressed when depleted of macroH2A. At the molecular level, we find that macroH2A is required for the interaction of repeat sequences with the nucleostructural protein lamin B1. Taken together, our results argue that a major function of macroH2A histone variants is to link nucleosome composition to higher-order chromatin architecture.

Spatio-temporal dynamics of ß-tubulin isotypes during the development of the sensory auditory organ in rat.

There are different ß-tubulin isoforms in microtubules of vertebrate tissues. However, their functional significance is still largely unknown. In the present study, we investigated the localization of five ß-tubulin isotypes (ß1-5) within the hearing organ during development in rat. By using confocal microscopy, we showed that with the exception of the ß3-tubulin isoform that was specific to nerve fibres, all the different ß-tubulin isoforms were mainly present in the supporting cells. Contrary to ß1-4-tubulins, we also found that the ß5-tubulin isoform appeared only at a key stage of the post-natal development in specific cell types (pillar cells and Deiters' cells). By using transmission electron microscopy, we revealed further that this developmental stage coincided with the formation of two separate bundles of microtubules from a unique one in these supporting cells. Together, these results suggest that the ß5-tubulin isoform might be involved in the generation of new microtubule bundles from a pre-existing one.

Large-scale annotated dataset for cochlear hair cell detection and classification.

Our sense of hearing is mediated by cochlear hair cells, of which there are two types organized in one row of inner hair cells and three rows of outer hair cells. Each cochlea contains 5-15 thousand terminally differentiated hair cells, and their survival is essential for hearing as they do not regenerate after insult. It is often desirable in hearing research to quantify the number of hair cells within cochlear samples, in both pathological conditions, and in response to treatment. Machine learning can be used to automate the quantification process but requires a vast and diverse dataset for effective training. In this study, we present a large collection of annotated cochlear hair-cell datasets, labeled with commonly used hair-cell markers and imaged using various fluorescence microscopy techniques. The collection includes samples from mouse, rat, guinea pig, pig, primate, and human cochlear tissue, from normal conditions and following in-vivo and in-vitro ototoxic drug application. The dataset includes over 107,000 hair cells which have been identified and annotated as either inner or outer hair cells. This dataset is the result of a collaborative effort from multiple laboratories and has been carefully curated to represent a variety of imaging techniques. With suggested usage parameters and a well-described annotation procedure, this collection can facilitate the development of generalizable cochlear hair-cell detection models or serve as a starting point for fine-tuning models for other analysis tasks. By providing this dataset, we aim to give other hearing research groups the opportunity to develop their own tools with which to analyze cochlear imaging data more fully, accurately, and with greater ease.

Large-scale annotated dataset for cochlear hair cell detection and classification.

Our sense of hearing is mediated by cochlear hair cells, localized within the sensory epithelium called the organ of Corti. There are two types of hair cells in the cochlea, which are organized in one row of inner hair cells and three rows of outer hair cells. Each cochlea contains a few thousands of hair cells, and their survival is essential for our perception of sound because they are terminally differentiated and do not regenerate after insult. It is often desirable in hearing research to quantify the number of hair cells within cochlear samples, in both pathological conditions, and in response to treatment. However, the sheer number of cells along the cochlea makes manual quantification impractical. Machine learning can be used to overcome this challenge by automating the quantification process but requires a vast and diverse dataset for effective training. In this study, we present a large collection of annotated cochlear hair-cell datasets, labeled with commonly used hair-cell markers and imaged using various fluorescence microscopy techniques. The collection includes samples from mouse, human, pig and guinea pig cochlear tissue, from normal conditions and following in-vivo and in-vitro ototoxic drug application. The dataset includes over 90'000 hair cells, all of which have been manually identified and annotated as one of two cell types: inner hair cells and outer hair cells. This dataset is the result of a collaborative effort from multiple laboratories and has been carefully curated to represent a variety of imaging techniques. With suggested usage parameters and a well-described annotation procedure, this collection can facilitate the development of generalizable cochlear hair cell detection models or serve as a starting point for fine-tuning models for other analysis tasks. By providing this dataset, we aim to supply other groups within the hearing research community with the opportunity to develop their own tools with which to analyze cochlear imaging data more fully, accurately, and with greater ease.

Intermediate Cells of Dual Embryonic Origin Follow a Basal to Apical Gradient of Ingression Into the Lateral Wall of the Cochlea.

Intermediate cells of the stria vascularis are neural crest derived melanocytes. They are essential for the establishment of the endocochlear potential in the inner ear, which allows mechanosensory hair cells to transduce sound into nerve impulses. Despite their importance for normal hearing, how these cells develop and migrate to their position in the lateral wall of the cochlea has not been studied. We find that as early as E10.5 some Schwann cell precursors in the VIIIth ganglion begin to express melanocyte specific markers while neural crest derived melanoblasts migrate into the otic vesicle. Intermediate cells of both melanoblast and Schwann cell precursor origin ingress into the lateral wall of the cochlea starting at around E15.5 following a basal to apical gradient during embryonic development, and continue to proliferate postnatally.

Congenital Deafness and Recent Advances Towards Restoring Hearing Loss.

Congenital hearing loss is the most common birth defect, estimated to affect 2-3 in every 1000 births. Currently there is no cure for hearing loss. Treatment options are limited to hearing aids for mild and moderate cases, and cochlear implants for severe and profound hearing loss. Here we provide a literature overview of the environmental and genetic causes of congenital hearing loss, common animal models and methods used for hearing research, as well as recent advances towards developing therapies to treat congenital deafness. © 2021 The Authors.

Transcriptomic analysis and ednrb expression in cochlear intermediate cells reveal developmental differences between inner ear and skin melanocytes.

In the inner ear, the neural crest gives rise to the glia of the VIII ganglion and two types of melanocytic cells: The pigmented cells of the vestibular system and intermediate cells of the stria vascularis. We analyzed the transcriptome of neonatal intermediate cells in an effort to better understand the development of the stria vascularis. We found that the expression of endothelin receptor B, which is essential for melanocyte development, persists in intermediate cells long after birth. In contrast, skin melanocytes rapidly downregulate the expression of EdnrB. Our findings suggest that endothelins might have co-opted new functions in the inner ear during evolution of the auditory organ.

MicroRNA-124 Regulates Cell Specification in the Cochlea through Modulation of Sfrp4/5.

The organ of Corti, the auditory organ of the mammalian inner ear, contains sensory hair cells and supporting cells that arise from a common sensory progenitor. The molecular bases allowing the specification of these progenitors remain elusive. In the present study, by combining microarray analyses with conditional deletion of Dicer in the developing inner ear, we identified that miR-124 controls cell fate in the developing organ of Corti. By targeting secreted frizzled-related protein 4 (Sfrp4) and Sfrp5, two inhibitors of the Wnt pathway, we showed that miR-124 controls the ß-catenin-dependent and also the PCP-related non-canonical Wnt pathways that contribute to HC differentiation and polarization in the organ of Corti. Thus, our work emphasizes the importance of miR-124 as an epigenetic safeguard that fine-tunes the expression of genes critical for cell patterning during cochlear differentiation.