3D in situ imaging of female reproductive tract reveals molecular signatures of fertilizing spermatozoa in mice
- IBT CAS and BIOCEV centre, Czech Republic
- Yale School of Medicine, United States
- Howard Hughes Medical Institute, Boston Children's Hospital, United States
- Baylor College of Medicine, United States
Abstract
Out of millions of ejaculated sperm, only a few reach the fertilization site in mammals. Flagellar Ca2+ signaling nanodomains, organized by multi-subunit CatSper calcium channel complexes, are pivotal for sperm migration in the female tract, implicating CatSper-dependent mechanisms in sperm selection. Here, using biochemical and pharmacological studies, we demonstrate that CatSper1 is an O-linked glycosylated protein, undergoing capacitation-induced processing dependent on Ca2+ and phosphorylation cascades. CatSper1 processing correlates with protein tyrosine phosphorylation (pY) development in sperm cells capacitated in vitro and in vivo. Using 3D in situ molecular imaging and ANN-based automatic detection of sperm distributed along the cleared female tract, we demonstrate that all spermatozoa past the UTJ possess intact CatSper1 signals. Together, we reveal that fertilizing mouse spermatozoa in situ are characterized by intact CatSper channel, lack of pY, and reacted acrosomes. These findings provide molecular insight into sperm selection for successful fertilization in the female reproductive tract.
Data availability
All data generated or analysed during this study are included in the manuscript, supplementary and source data files.
Article and author information
Author details
Funding
National Institutes of Health (R01HD096745)
- Jean-Ju Chung
Yale School of Medicine (Start-up funds)
- Jean-Ju Chung
Yale University (a Yale Goodman-Gilman ScholarAward-2015)
- Jean-Ju Chung
Male Contraceptive Initiative (Postdoctoral fellowship)
- Jae Yeon Hwang
Czech Science Foundation (GJ20-17403Y)
- Lukas Ded
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Ethics
Animal experimentation: Animal experimentation: This study was performed in strict accordance with the recommendations in the Guide for the Care and Use of Laboratory Animals of the National Institutes of Health. All the mice were treated in accordance with guidelines approved by Yale (20079) Animal Care and Use Committees (IACUC).
Copyright
© 2020, Ded 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.
Metrics
-
- 4,788
- views
-
- 595
- downloads
-
- 36
- citations
Views, downloads and citations are aggregated across all versions of this paper published by eLife.
Download links
A two-part list of links to download the article, or parts of the article, in various formats.
Downloads (link to download the article as PDF)
Open citations (links to open the citations from this article in various online reference manager services)
Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)
3D in situ imaging of female reproductive tract reveals molecular signatures of fertilizing spermatozoa in mice
eLife 9:e62043.
https://doi.org/10.7554/eLife.62043
Further reading
-
- Cell Biology
- Developmental Biology
A study in mice reveals key interactions between proteins involved in fibroblast growth factor signaling and how they contribute to distinct stages of eye lens development.
-
- Cell Biology
- Evolutionary Biology
Maintenance of rod-shape in bacterial cells depends on the actin-like protein MreB. Deletion of mreB from Pseudomonas fluorescens SBW25 results in viable spherical cells of variable volume and reduced fitness. Using a combination of time-resolved microscopy and biochemical assay of peptidoglycan synthesis, we show that reduced fitness is a consequence of perturbed cell size homeostasis that arises primarily from differential growth of daughter cells. A 1000-generation selection experiment resulted in rapid restoration of fitness with derived cells retaining spherical shape. Mutations in the peptidoglycan synthesis protein Pbp1A were identified as the main route for evolutionary rescue with genetic reconstructions demonstrating causality. Compensatory pbp1A mutations that targeted transpeptidase activity enhanced homogeneity of cell wall synthesis on lateral surfaces and restored cell size homeostasis. Mechanistic explanations require enhanced understanding of why deletion of mreB causes heterogeneity in cell wall synthesis. We conclude by presenting two testable hypotheses, one of which posits that heterogeneity stems from non-functional cell wall synthesis machinery, while the second posits that the machinery is functional, albeit stalled. Overall, our data provide support for the second hypothesis and draw attention to the importance of balance between transpeptidase and glycosyltransferase functions of peptidoglycan building enzymes for cell shape determination.
