Neurodevelopmental axonal pathfinding plays a central role in correct brain wiring and subsequent cognitive abilities. Within the growth cone, various intracellular effectors transduce axonal guidance signals by remodeling the cytoskeleton. Semaphorin-3E (Sema3E) is a guidance cue implicated in development of the fornix, a neuronal tract connecting the hippocampus to the hypothalamus. Microtubule-Associated Protein 6 (MAP6) has been shown to be involved in the Sema3E growth-promoting signaling pathway. In this study, we identified the Collapsin Response Mediator Protein 4 (CRMP4) as a MAP6 partner and a crucial effector in Sema3E growth-promoting activity. CRMP4-KO mice displayed abnormal fornix development reminiscent of that observed in Sema3E-KO mice. CRMP4 was shown to interact with the Sema3E tripartite receptor complex within Detergent-Resistant Membrane (DRM) domains, and DRM domain integrity was required to transduce Sema3E signaling through the Akt/GSK3 pathway. Finally, we showed that the cytoskeleton-binding domain of CRMP4 is required for Sema3E's growth-promoting activity, suggesting that CRMP4 plays a role at the interface between Sema3E receptors, located in DRM domains, and the cytoskeleton network. As the fornix is affected in many psychiatric diseases, such as schizophrenia, our results provide new insights to better understand the neurodevelopmental components of these diseases.
All data generated or analysed during this study are included in the manuscript and supporting files. Source data files have been provided for Figure 2, Figure 4, Figure 5, Figure 6 , Figure 7, Figure 8, Figure 9, Figure 10 and supplementary File 1
- Christophe Bosc
- Annie Andrieux
The funder had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Animal experimentation: The study protocol was approved by the local animal welfare committee (Comité Local GIN, C2EA-04 - APAFIS number 8303-2016060110523424) and complied with EU guidelines (directive 2010/63/EU). Every precaution was taken to minimize the number of animals used and stress to animals during experiments.
- Fadel Tissir, Université Catholique de Louvain, Belgium
- Received: May 14, 2021
- Accepted: December 2, 2021
- Accepted Manuscript published: December 3, 2021 (version 1)
- Accepted Manuscript updated: December 6, 2021 (version 2)
- Accepted Manuscript updated: December 6, 2021 (version 3)
- Accepted Manuscript updated: December 13, 2021 (version 4)
- Version of Record published: December 17, 2021 (version 5)
- Version of Record updated: December 20, 2021 (version 6)
© 2021, Boulan 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.