Identification and characterization of mouse otic sensory lineage genes

Abstract

Vertebrate embryogenesis gives rise to all cell types of an organism through the development of many unique lineages derived from the three primordial germ layers. The otic sensory lineage arises from the otic vesicle, a structure formed through invagination of placodal non-neural ectoderm. This developmental lineage possesses unique differentiation potential, giving rise to otic sensory cell populations including hair cells, supporting cells, and ganglion neurons of the auditory and vestibular organs. Here we present a systematic approach to identify transcriptional features that distinguish the otic sensory lineage (from early otic progenitors to otic sensory populations) from other major lineages of vertebrate development. We used a microarray approach to analyze otic sensory lineage populations including microdissected otic vesicles (embryonic day 10.5) as well as isolated neonatal cochlear hair cells and supporting cells at postnatal day 3. Non-otic tissue samples including periotic tissues and whole embryos with otic regions removed were used as reference populations to evaluate otic specificity. Otic populations shared transcriptome-wide correlations in expression profiles that distinguish members of this lineage from non-otic populations. We further analyzed the microarray data using comparative and dimension reduction methods to identify individual genes that are specifically expressed in the otic sensory lineage. This analysis identified and ranked top otic sensory lineage-specific transcripts including Fbxo2, Col9a2, and Oc90, and additional novel otic lineage markers. To validate these results we performed expression analysis on select genes using immunohistochemistry and in situ hybridization. Fbxo2 showed the most striking pattern of specificity to the otic sensory lineage, including robust expression in the early otic vesicle and sustained expression in prosensory progenitors and auditory and vestibular hair cells and supporting cells.

Neural stem/progenitor cells are activated during tail regeneration in the leopard gecko (Eublepharis macularius)

“…TUJ1 (Beta‐III‐Tubulin; Millipore, USA, Cat#MAB5564, RRID AB_11212768) is a mouse monoclonal antibody raised against purified immunoglobulins (manufacturer's information). A single ∼50 kDa band, the expected molecular weight (manufacturer's information), was detected on Western blots of gecko brain and spinal cord homogenates (data not shown) TUJ1 immunostaining is exclusive to neuronal‐like cells of the spinal cord and matches the staining pattern observed in mice (Hartman, Durruthy‐Durruthy, Laske, Losorelli, & Heller, ).This antibody also stains rat neuronal cells in vitro (Takaki et al, ).…”

“…Using immunohistochemistry, SOX9 immunostaining was restricted to the nuclei of chondrocytes and ependymal cells of the spinal cord (McLean & Vickaryous, 2011).TUJ1 (Beta-III-Tubulin; Millipore, USA, Cat#MAB5564, RRID AB_11212768) is a mouse monoclonal antibody raised against purified immunoglobulins (manufacturer's information). A single 50 kDa band, the expected molecular weight (manufacturer's information), was detected on Western blots of gecko brain and spinal cord homogenates (data not shown) TUJ1 immunostaining is exclusive to neuronal-like cells of the spinal cord and matches the staining pattern observed in mice(Hartman, Durruthy-Durruthy, Laske, Losorelli, & Heller, 2015).This antibody also stains rat neuronal cells in vitro(Takaki et al, 2012).Vimentin (Developmental Studies Hybridoma Bank, USA, Cat# H5, RRID AB_528506) is a mouse monoclonal antibody raised against vimentin isolated from chick optic tectum (manufacturer's information). A single 52 kDa band, the expected molecular weight (manufacturer's information), was detected on Western blots of gecko brain and spinal cord homogenates (data not shown).…”

SoxC transcription factors are essential for the development of the inner ear

“…With a simple architecture and just under 4,000 hair cells in an adult animal (14), the sensory epithelium of the utricle-the macularepresents a useful model system. Although gene expression has been characterized in early otic development (15,16) and in the neonatal organ of Corti (17,18), corresponding data are lacking for the developing utricle.…”

Single-cell analysis delineates a trajectory toward the human early otic lineage

“…Without a complex 3D microenvironment (19,21), proper sensory hair cell differentiation only sporadically occurs in cocultures with primary embryonic otic cells (8). Whereas guidance protocols for mouse otic lineage cells are continuously evolving through novel tools such as cellular reprogramming (22) or highly specific novel otic lineage markers (23), generation of human hair cell-like cells remains perplexingly difficult. The lack of specific human otic lineage markers impedes identification and isolation of human otic lineage cells.…”