Characterization of transcriptomes of cochlear inner and outer hair cells

Abstract

Inner hair cells (IHCs) and outer hair cells (OHCs) are the two types of sensory receptor cells that are critical for hearing in the mammalian cochlea. IHCs and OHCs have different morphology and function. The genetic mechanisms that define their morphological and functional specializations are essentially unknown. The transcriptome reflects the genes that are being actively expressed in a cell and holds the key to understanding the molecular mechanisms of the biological properties of the cell. Using DNA microarray, we examined the transcriptome of 2000 individually collected IHCs and OHCs from adult mouse cochleae. We show that 16,647 and 17,711 transcripts are expressed in IHCs and OHCs, respectively. Of those genes, ∼73% are known genes, 22% are uncharacterized sequences, and 5.0% are noncoding RNAs in both populations. A total of 16,117 transcripts are expressed in both populations. Uniquely and differentially expressed genes account for <15% of all genes in either cell type. The top 10 differentially expressed genes include Slc17a8, Dnajc5b, Slc1a3, Atp2a3, Osbpl6, Slc7a14, Bcl2, Bin1, Prkd1, and Map4k4 in IHCs and Slc26a5, C1ql1, Strc, Dnm3, Plbd1, Lbh, Olfm1, Plce1, Tectb, and Ankrd22 in OHCs. We analyzed commonly and differentially expressed genes with the focus on genes related to hair cell specializations in the apical, basolateral, and synaptic membranes. Eighty-three percent of the known deafness-related genes are expressed in hair cells. We also analyzed genes involved in cell-cycle regulation. Our dataset holds an extraordinary trove of information about the molecular mechanisms underlying hair cell morphology, function, pathology, and cell-cycle control.

Keywords: DNA microarray; inner hair cells; mouse; outer hair cells; transcriptome.

Figure 1.

Micrographs of isolated hair cells and supporting cells and a pickup pipette used to collect isolated hair cells. A, Images of a pickup pipette before and after an isolated OHC was drawn into the pipette. B, Images (from left to right) of an isolated IHC, OHC, pillar cell, and Deiters' cell from adult mice. Scale bar, 5 μm for all images in panel B.

Figure 2.

Expression levels of the top 200 genes in IHCs in descending order. The expression value of the same genes in OHCs is also presented. Numbers in red signify the abundance rank of the genes in IHCs; the numbers in blue on the right represent the ranking of the same genes in OHCs. In this and all subsequent figures, ENSMUST identification numbers are truncated to show the last six digits.

Figure 3.

Expression levels of the top 200 genes in OHCs. The expression value of the same genes in IHCs is also presented for comparison. Numbers in blue signify the abundance rank of the genes in OHCs; the numbers in red on the right represent the ranking of the same genes in IHCs.

Figure 4.

Differentially expressed genes in IHCs and OHCs. A, All differentially expressed genes in IHCs and OHCs. The red line represents the expression level of 18,240 transcripts of IHCs; each blue dot represents the expression level of the same transcripts of OHCs. Genes that are differentially expressed in OHCs are above the red line; genes that are differentially expressed in IHCs are below the red line. Uniquely expressed genes in IHCs and OHCs are highlighted in the light purple and light green areas. Some representative genes that are differentially expressed in IHCs and OHCs are marked by purple dots. B, The 150 most differentially expressed genes in IHCs. The numerical values in red on the right represent the fold difference in expression for IHCs versus OHCs. C, The 150 most differentially expressed genes in OHCs. The numerical values in blue on the right represent the fold difference in expression for OHCs versus IHCs.

Figure 5.

The top 150 uniquely expressed genes in IHCs and OHCs. Uniquely expressed genes are those whose expression levels were below background in only one cell type. A, The expression levels of the top 150 uniquely expressed genes in IHCs. B, The expression levels of the top 150 uniquely expressed genes in OHCs.

Figure 6.

Genes related to stereocilia structure, ion channels, neurotransmitter vesicle release, and ACh receptor. A, Comparison of expression levels for genes that encode proteins associated with stereocilia bundles. Numerical values to the right represent the amount of protein detected in stereocilia by Shin et al. (2013). B, Comparison of expression levels for genes associated with ion channels, neurotransmitter vesicle transport and release, and ACh receptors.

Figure 7.

Genes regulating cell cycle and genes related to deafness. A, Expression levels of 85 genes that are important for cell-cycle regulation. B, Comparison of expression levels for 101 known deafness genes. The blue line in each panel marks the baseline for expression. Genes whose expression levels fall below the baseline are not considered to be expressed.