Abstract
Spermatozoa are polarized cells with a head and a flagellum joined together by the connecting piece. Flagellum integrity is critical for normal sperm function, and flagellum defects consistently lead to male infertility. Multiple morphological abnormalities of the flagella (MMAF) is a distinct sperm phenotype consistently leading to male infertility due to a reduced or absent sperm motility associated with severe morphological and ultrastructural flagellum defects. Despite numerous genes recently described to be recurrently associated with MMAF, more than half of the cases analyzed remain unresolved, suggesting that many yet uncharacterized gene defects account for this phenotype. By performing a retrospective exome analysis of the unsolved cases from our initial cohort of 167 infertile men with a MMAF phenotype, we identified one individual carrying a homozygous frameshift variant in CFAP206, a gene encoding a microtubule-docking adapter for radial spoke and inner dynein arm. Immunostaining experiments in the patient’s sperm cells demonstrated the absence of WDR66 and RSPH1 proteins suggesting severe radial spokes and calmodulin and spoke-associated complex defects. Using the CRISPR–Cas9 technique, we generated homozygous Cfap206 knockout (KO) mice which presented with male infertility due to functional, structural and ultrastructural sperm flagellum defects associated with a very low rate of embryo development using ICSI. Overall, we showed that CFAP206 is essential for normal sperm flagellum structure and function in human and mouse and that bi-allelic mutations in CFAP206 cause male infertility in man and mouse by inducing morphological and functional defects of the sperm flagellum that may also cause ICSI failures.
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Data availability
The datasets generated during and/or analyzed during the current study are available from the corresponding author on reasonable request. Accession numbers: The CFAP206 variant reported in this manuscript is accessible in ClinVar with the submission number SUB9294549.
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Acknowledgements
We thank the patient for their participation in this study as well as all the referring physicians.
Funding
This work was mainly supported by the following grants by the Agence Nationale de la Recherche (ANR) MASFLAGELLA (ANR-14-CE15-0002) and FLAGEL-OME (ANR-19-CE17-0014). This work was also supported by the National Key R&D Program of China [Grant number 2019YFC1005106]; Supported by the Non-profit Central Research Institute Fund of Chinese Academy of Medical Sciences (2019PT310002; the National Natural Science Foundation of China [Grant numbers 81901541 and 81971441].
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All authors contributed to the study conception and design. CCo, YXC, PFR, FZ, XJH and XA designed and supervised the study. AA-Y, VS recruited the patients, performed clinical analysis and characterization. NTM, ZEK, PFR, TC and CCo performed exome and bioinformatics analysis. GM, JB, CCa, MB and TC performed patient’s experimental work. HBL and QSS provided Cfap206 knockout mice. BS, MRL and QSS performed animal’s experimental work. QSS and DL performed ICSI of mice. GM, CCA, KKL, FBT and XJH performed data illustrations. XA, FZ, PFR, AT, MW, CA, HW and CCo analyzed the data. XA, GM, QSS and CCo wrote the manuscript. The first draft of the manuscript was written by XA, GM and CCo, and all authors commented on previous versions of the manuscript. All authors read and approved the final manuscript.
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The study was approved by local ethics committees, and samples were then stored in the CRB Germethèque (certification under ISO-9001 and NF-S 96-900) following a standardized procedure or were part of the Fertithèque collection declared to the French Ministry of health (DC-2015-2580) and the French Data Protection Authority (DR-2016-392).
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All animal procedures were run according to the Chinese guidelines on the use of animals in scientific investigations with the approval of the animal Ethics Committee at the First Affiliated Hospital of Anhui Medical University (P2020-12-36).
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ClinVar, https://www.ncbi.nlm.nih.gov/clinvar/
gnomAD Browser, http://gnomad.broadinstitute.org
GTEx, https://gtexportal.org
Uniprot, https://www.uniprot.org/
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Shen, Q., Martinez, G., Liu, H. et al. Bi-allelic truncating variants in CFAP206 cause male infertility in human and mouse. Hum Genet 140, 1367–1377 (2021). https://doi.org/10.1007/s00439-021-02313-z
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DOI: https://doi.org/10.1007/s00439-021-02313-z