Expression profiling of murine inner-ear hair cells
Ontology highlight
ABSTRACT: Murine auditory and vestibular epithelia were separated into hair-cells (HCs), epithelial non-sensory cells (ENSCs) and non-epithelial cells (NECs) by flow cytometry. Gene expression levels were recorded in independent triplicates from the sorted cells using Illumina whole genome expression microarrays
Project description:The mammalian inner ear subserves auditory and vestibular sensations via highly specialized cells and proteins. We show that sensory hair cells (HCs) employ hundreds of uniquely or highly expressed proteins for processes involved in transducing mechanical inputs, stimulating sensory neurons, and maintaining structure and function of these post-mitotic cells. Our proteomic analysis of purified HCs extends the existing HC transcriptome, revealing undetected gene products and isoform-specific protein expression. Comparison with mouse and human databases of genetic auditory/vestibular impairments confirms the critical role of the HC proteome for normal inner ear function, providing a cell-specific pool of candidates for novel, important HC genes. Several proteins identified exclusively in HCs by proteomics and by immunohistochemistry map to human genetic deafness loci, potentially representing new deafness genes.
Project description:To understand the basic biological properties of inner ear hair cells (HCs) from non-mammalian vertebrates, we examined the transcriptome of adult zebrafish auditory and vestibular HCs. GFP-labeled HCs were isolated from inner-ear sensory epithelia of a pou4f3 promoter-driven GAP-GFP line of transgenic zebrafish. One thousand HCs and 1,000 non-sensory surrounding cells (nsSCs) were separately collected for each biological replicate, using the suction pipette technique. RNA sequencing of three biological replicates for the two cell types was performed. The resulting sequenced reads were mapped. Comparisons between HCs and nsSCs allow identification of enriched genes in HCs, which may underlie HC specialization. Our dataset provides an extensive resource for understanding the molecular mechanisms underlying morphology, function, and pathology of adult zebrafish HCs. It also establishes a framework for future characterization of the genes expressed in HCs and for the study of HC evolution.
Project description:The inner ear continues to grow and develop until the auditory and vestibular systems reach full maturity and all of the genes involved in this process have yet to be identified. Previous gene based analysis have primarily focused on the early developmental stages following induction and initial formation of the inner ear. The aim of this study is to identify new candidate genes for inner ear development. Microarrays were used to produce expression profiles from larval stages 56,57,58 of the Xenopus laevis inner ear. The data produced from this work represent an annotated resource that can be utilized by the Xenopus community to provide candidates for further functional analysis. Xenopus inner ears were isolated from larval animals for RNA extraction and hybridization to Affymetrix GeneChip microarrays.
Project description:The inner ear continues to grow and develop until the auditory and vestibular systems reach full maturity and all of the genes involved in this process have yet to be identified. Previous gene based analysis have primarily focused on the early developmental stages following induction and initial formation of the inner ear. The aim of this study is to identify new candidate genes for inner ear development. Microarrays were used to produce expression profiles from larval stages 56,57,58 of the Xenopus laevis inner ear. The data produced from this work represent an annotated resource that can be utilized by the Xenopus community to provide candidates for further functional analysis.
Project description:The inner ear continues to grow and develop until the auditory and vestibular systems reach full maturity and all of the genes involved in this process have yet to be identified. Previous gene based analysis have primarily focused on the early developmental stages following induction and initial formation of the inner ear. The aim of this study is to identify new candidate genes for inner ear development. Microarrays were used to produce expression profiles from larval stages 50,51,52 of the Xenopus laevis inner ear. The data produced from this work represent an annotated resource that can be utilized by the Xenopus community to provide candidates for further functional analysis.
Project description:The genes involved in inner ear development have yet to be fully characterized. Previous gene-based analyses have primarily focused on the early developmental stages following induction and initial formation of the inner ear. The inner ear continues to grow and develop until the auditory and vestibular systems reach full maturity; all of the genes involved in this process have yet to be identified. The aim of this study is to identify additional candidate genes for inner ear development. Microarrays were used to produce expression profiles from the post-metamorphic juvenile stage of the Xenopus laevis inner ear.
Project description:In the inner ear, cochlear and vestibular sensory epithelia utilize grossly similar cell types to transduce different stimuli: sound and acceleration. Each individual sensory epithelium is composed of highly heterogeneous populations of cells based on physiological and anatomical criteria. However, limited numbers of each cell type have impeded transcriptional characterization. Here we generated transcriptomes for 301 single cells from the utricular and cochlear sensory epithelia of newborn mice to circumvent this challenge. Cluster analysis indicates distinct profiles for each of the major sensory epithelial cell types, as well as less distinct subpopulations. Asynchrony within utricles allows reconstruction of the temporal progression of cell-type specific differentiation and suggests possible plasticity among cells at the sensory-nonsensory boundary. Comparisons of cell types from utricles and cochleae demonstrate divergence between auditory and vestibular cells despite a common origin. These results provide significant insights into the developmental processes that form unique inner ear cell types.
Project description:Sensorineural hearing loss affects the majority of the elderly population. Mammalian hair cells (HC) do not regenerate and current stem cell and gene delivery protocols result only in immature hair cells like-cells. For this reason, characterization of the transcriptional cascades that lead to development and survival of inner ear HC is essential for designing molecular-based treatments for deafness. We employed a cell type-specific approach to analyze the transcriptomes of the mouse early postnatal auditory and vestibular sensory epithelia and of hair cells derived from zebrafish model.
Project description:We have employed a novel approach for the identification of functionally important microRNA (miRNA)-target interactions using integrated miRNA, transcriptome and proteome profiles with advanced in silico analysis. By looking at both the transcript and protein levels of expression, a thorough coverage of miRNA regulation was obtained. Microdissected auditory and vestibular sensory epithelia were used as the model system, thus being the first time such a comparison was carried out in a neuroepithelial system. Moreover, this is one of only a few studies employing proteome screening for the identification of miRNA targets. Notably, this approach can be employed for the study of other tissues and organs. We detected the expression of 157 miRNAs in the inner ear sensory epithelia, with 53 miRNAs differentially expressed between the cochlea and vestibule. By searching for enrichment and depletion of miRNA targets in the transcript and protein datasets with a reciprocal or similar expression, respectively, as the regulatory miRNA, we identified functionally important miRNAs. Finally, the interaction between miR-135b and PSIP1-P75, a transcriptional coacitvator previously unknown in the inner ear, was identified and validated experimentally. We suggest that miR-135b may serve as a cellular effector, involved in regulating some of the differences between the cochlear and vestibular hair cells. We investigated the mRNA expression profile of the cochlear and vestibular sensory epithelia from inner ears of postnatal day 2 mice using the Affymetrix GeneChip® 430 2Mouse Genome array. Cochlear and vestibular sensory epithelia were dissected from wild type C3H mice and collected separately. The vestibular epithelia consisted of the saccule, utricle and the lateral and anterior cristae. Both the cochlear and vestibular sensory epithelia were dissected with their underlying mesenchyma. Altogether three pools, three biological replicates, of each tissue type were collected consisting of cochlear or vestibular sensory epithelia dissected from 10 to 12 inner ears.
Project description:The correlation between expression levels of mRNA and protein in mammals is relatively low, with a Pearson correlation coefficient of ~0.40. Post transcriptional regulation contributes to this low correlation. Across taxa, it was demonstrated that translation efficiency, i.e. the protein-to-mRNA ratio in steady state, varies between species in a direction that buffers the protein levels from changes in the transcript abundance. Evidence for this behavior in tissues is sparse. We asked whether this phenomenon is evident in our auditory system data, and on previously obtained proteomic and transcriptomic data from different tissue datasets.