This study provides transcriptomic characterization of the cells of the crista ampullaris, sensory structures at the base of the semicircular canals that are critical for vestibular function. We performed single cell RNA-seq on ampullae microdissected from E16, E18, P3 and P7 mice. Cluster analysis identified the hair cells, support cells and glia of the crista as well as dark cells and other nonsensory epithelial cells of the ampulla, mesenchymal cells, vascular cells, macrophages and melanocytes. Cluster-specific expression of genes predicted their spatially restricted domains of gene expression in the crista and ampulla. Analysis of cellular proportions across developmental time showed dynamics in cellular composition. The new cell types revealed by single cell RNA-seq could be important for understanding crista function and the markers identified in this study will enable the examination of their dynamics during development and disease.
All RNA-Seq data has been ave been deposited in NCBI's Gene Expression Omnibus (Edgar et al., 2002) and are accessible through GEO Series accession number# GSE168901.
Single-cell Transcriptomic Analysis of the Mouse Crista AmpullarisNCBI Gene Expression Omnibus, GSE168901.
Characterization of cochlear development at the single cell levelNCBI Gene Expression Omnibus, GSE137299.
- Olivia Bermingham-McDonogh
- Brent A Wilkerson
- Brent A Wilkerson
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Animal experimentation: All animal work was approved by the Institutional Animal Care and Use committee under protocol number 3123-01 and conforms to the recommendations of the NIH for animal care and use. Every effort was made to minimize any distress to the mice. Euthanasia of mice was in accordance with the AVMA guidelines.
- Tanya T Whitfield, University of Sheffield, United Kingdom
© 2021, Wilkerson et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
Imaging experiments reveal the complex and dynamic nature of the transcriptional hubs associated with Notch signaling.
Cylicins are testis-specific proteins, which are exclusively expressed during spermiogenesis. In mice and humans, two Cylicins, the gonosomal X-linked Cylicin 1 (Cylc1/CYLC1) and the autosomal Cylicin 2 (Cylc2/CYLC2) genes, have been identified. Cylicins are cytoskeletal proteins with an overall positive charge due to lysine-rich repeats. While Cylicins have been localized in the acrosomal region of round spermatids, they resemble a major component of the calyx within the perinuclear theca at the posterior part of mature sperm nuclei. However, the role of Cylicins during spermiogenesis has not yet been investigated. Here, we applied CRISPR/Cas9-mediated gene editing in zygotes to establish Cylc1- and Cylc2-deficient mouse lines as a model to study the function of these proteins. Cylc1 deficiency resulted in male subfertility, whereas Cylc2-/-, Cylc1-/yCylc2+/-, and Cylc1-/yCylc2-/- males were infertile. Phenotypical characterization revealed that loss of Cylicins prevents proper calyx assembly during spermiogenesis. This results in decreased epididymal sperm counts, impaired shedding of excess cytoplasm, and severe structural malformations, ultimately resulting in impaired sperm motility. Furthermore, exome sequencing identified an infertile man with a hemizygous variant in CYLC1 and a heterozygous variant in CYLC2, displaying morphological abnormalities of the sperm including the absence of the acrosome. Thus, our study highlights the relevance and importance of Cylicins for spermiogenic remodeling and male fertility in human and mouse, and provides the basis for further studies on unraveling the complex molecular interactions between perinuclear theca proteins required during spermiogenesis.