Transcription Factors Expressed in Mouse Cochlear Inner and Outer Hair Cells

Abstract

Regulation of gene expression is essential to determining the functional complexity and morphological diversity seen among different cells. Transcriptional regulation is a crucial step in gene expression regulation because the genetic information is directly read from DNA by sequence-specific transcription factors (TFs). Although several mouse TF databases created from genome sequences and transcriptomes are available, a cell type-specific TF database from any normal cell populations is still lacking. We identify cell type-specific TF genes expressed in cochlear inner hair cells (IHCs) and outer hair cells (OHCs) using hair cell-specific transcriptomes from adult mice. IHCs and OHCs are the two types of sensory receptor cells in the mammalian cochlea. We show that 1,563 and 1,616 TF genes are respectively expressed in IHCs and OHCs among 2,230 putative mouse TF genes. While 1,536 are commonly expressed in both populations, 73 genes are differentially expressed (with at least a twofold difference) in IHCs and 13 are differentially expressed in OHCs. Our datasets represent the first cell type-specific TF databases for two populations of sensory receptor cells and are key informational resources for understanding the molecular mechanism underlying the biological properties and phenotypical differences of these cells.

Fig 1. Expression levels of the top 200 TF genes in adult IHCs (in red) in descending order.

For comparison, the expression value of the same genes in adult OHCs (in blue) is also presented. Color-coded numbers on the right side of each panel represents the abundance ranking of the each TF among 16,647 transcripts considered to be expressed in adult IHCs.

Fig 2. Expression levels of the top 200 TF genes in adult OHCs (in blue) in descending order.

For comparison, the expression value of the same genes in adult IHCs (in red) is also presented. Color-coded numbers on the right side of each panel represents the abundance ranking of the each TF among 17,711 transcripts considered to be expressed in adult OHCs.

Fig 3. Differentially and uniquely expressed TF genes in adult IHCs and OHCs.

(A) All TF genes in IHCs and OHCs. The red line represents the expression level of all individual TF genes of IHCs while each blue dot represents the expression level of the same TF genes in OHCs. TF genes that are differentially expressed in OHCs are above the red line while genes that are differentially expressed in IHCs are below the red line. (B) Differentially expressed IHC TF genes with reference to the expression of same genes in OHCs (in Log₂). (C) Differentially expressed adult OHC TF genes with reference to the expression of the same genes in IHCs. Only those whose expression levels differ by at least one Log₂-fold between the two populations of cells are shown in B and C.

Fig 4. Expression levels of the top 200 TF genes expressed in P7 HCs.

The expression of the same TF genes between E16 and P4 is also plotted for comparison. Gene expression profiles at different ages are color-coded. 55 TF genes that are expressed in both P7 and adult HCs are marked by red asterisks.

Fig 5. The top 100 TF genes that are up- and down-regulated the expression levels between E16 and P7 were compared with P7 as reference.

Fig 6. The expression levels of some TFs that are critical for hair cell and/or supporting cell development.

(A) TFs in adult IHCs and OHCs. (B) TFs in nascent hair cells.

Fig 7. Validation of gene expression by qPCR and immunocytochemistry.

(A) Comparison of seven TF genes expressed in adult IHCs and OHCs using qPCR. Note that the ordinate is the inverse of the number of PCR cycles necessary to reach fluorescence threshold. So even a small change (such as 0.01) reflects a large difference. The asterisk represents statistical significance (p < 0.001, Student’s t-test) between each pair. (B) Fluorescence image of expression of LBH in the adult organ of Corti. (C) Optical sectioning of the organ of Corti using confocal microscopy. Nuclei were labelled with DAPI. Bar represents 10 μm.