[1] [1] Afzelius, B. A. Cilia-related diseases.J. Pathol.204, 470–477(2004).

[2] [2] Reiter, J. F. & Leroux, M. R. Genes and molecular pathways underpinning ciliopathies.Nat. Rev. Mol. Cell Biol.18, 533–547(2017).

[3] [3] Hildebrandt, F., Benzing, T. & Katsanis, N. Ciliopathies.N. Engl. J. Med.364, 1533–1543(2011).

[4] [4] Viswanadha, R., Sale, W. S. & Porter, M. E. Ciliary motility: regulation of axonemal dynein motors.Cold Spring Harb. Perspect. Biol.9, a018325(2017).

[5] [5] Ishikawa, T. Axoneme structure from motile cilia.Cold Spring Harb. Perspect. Biol.9, a018325(2017).

[6] [6] Omoto, C. K. et al. Rotation of the central pair microtubules in eukaryotic flagella.Mol. Biol. Cell10, 1–4(1999).

[7] [7] Oda, T., Yanagisawa, H., Yagi, T. & Kikkawa, M. Mechanosignaling between central apparatus and radial spokes controls axonemal dynein activity.J. Cell Biol.204, 807–819(2014).

[8] [8] Smith, E. F. & Yang, P. F. The radial spokes and central apparatus: mechanochemical transducers that regulate flagellar motility.Cell Motil. Cytoskel.57, 8–17(2004).

[9] [9] Yoshimura, M. & Shingyoji, C. Effects of the central pair apparatus on microtubule sliding velocity in sea urchin sperm flagella.Cell Struct. Funct.24, 43–54(1999).

[10] [10] Carbajal-Gonzlez, B. I. et al. Conserved structural motifs in the central pair complex of eukaryotic flagella.Cytoskeleton70, 101–120(2013).

[11] [11] Han, L. et al. Cryo-EM structure of an active central apparatus.Nat. Struct. Mol. Biol.29, 472–482(2022).

[12] [12] Gui, M., Wang, X., Dutcher, S. K., Brown, A. & Zhang, R. Ciliary central apparatus structure reveals mechanisms of microtubule patterning.Nat. Struct. Mol. Biol.29, 483–492(2022).

[13] [13] Teves, M. E., Nagarkatti-Gude, D. R., Zhang, Z. B. & Strauss, J. F. Mammalian axoneme central pair complex proteins: Broader roles revealed by gene knockout phenotypes.Cytoskeleton73, 3–22(2016).

[14] [14] Chen, Z. et al. In situ cryo-electron tomography reveals the asymmetric architecture of mammalian sperm axonemes.Nat. Struct. Mol. Biol.30, 360–369(2023).

[15] [15] Leung, M. R. et al. The multi-scale architecture of mammalian sperm flagella and implications for ciliary motility.EMBO J.40, e107410(2021).

[16] [16] Chen, Z. et al. De novo protein identification in mammalian sperm using in situ cryoelectron tomography and AlphaFold2 docking.Cell186, 5041–5053(2023).

[17] [17] Tai, L., Yin, G., Huang, X., Sun, F. & Zhu, Y. In-cell structural insight into the stability of sperm microtubule doublet.Cell Discov.9, 116(2023).

[18] [18] Skerrett-Byrne, D. A. et al. Global profiling of the proteomic changes associated with the post-testicular maturation of mouse spermatozoa.Cell Rep.41, 111655(2022).

[19] [19] Gao, J. et al. DomainFit: Identification of protein domains in cryo-EM maps at intermediate resolutionusing AlphaFold2-predictedmodels.Structure32, 1248–1259(2024).

[20] [20] Ponting, C. P. A novel domain suggests a ciliary function for ASPM, a brain size determining gene.Bioinformatics22, 1031–1035(2006).

[21] [21] Fu, G. et al. Structural organization of the C1a-e-c supercomplex within the ciliary central apparatus.J. Cell Biol.218, 4236–4251(2019).

[22] [22] Walensky, L. D., Roskams, A. J., Lefkowitz, R. J., Snyder, S. H. & Ronnett, G. V. Odorant receptors and desensitization proteins colocalize in mammalian sperm.Mol. Med.1, 130–141(1995).

[23] [23] Smith, E. F. & Lefebvre, P. A. PF20 gene product contains WD repeats and localizes to the intermicrotubule bridges in Chlamydomonas flagella.Mol. Biol. Cell8, 455–467(1997).

[24] [24] Teves, M. E. et al. Sperm-associated antigen-17 gene is essential for motile cilia function and neonatal survival.Am. J. Respir. Cell Mol. Biol.48, 765–772(2013).

[25] [25] Leung, M. R. et al. Structural diversity of axonemes across mammalian motile cilia.Nature637, 1170–1177(2025).

[26] [26] Tanaka-Hayashi, A. et al. Is D-aspartate produced by glutamic-oxaloacetic transaminase-1 like 1 (Got1l1): a putative aspartate racemase?Amino Acids47, 79–86(2015).

[27] [27] Tomita, K. et al. The effect of D-aspartate on spermatogenesis in mouse testis.Biol. Reprod.94, 30(2016).

[28] [28] Tegha-Dunghu, J. et al. MAP1S controls microtubule stability throughout the cell cycle in human cells.J. Cell Sci.127, 5007–5013(2014).

[29] [29] Cheung, S., Xie, P., Rosenwaks, Z. & Palermo, G. D. Profiling the male germline genome to unravel its reproductive potential.Fertil. Steril.119, 196–206(2023).

[30] [30] Gui, M. et al. SPACA9 is a lumenal protein of human ciliary singlet and doublet microtubules.Proc. Natl. Acad. Sci. USA119, e2207605119(2022).

[31] [31] Gui, M. et al. Structures of radial spokes and associated complexes important for ciliary motility.Nat. Struct. Mol. Biol.28, 29–37(2021).

[32] [32] Grossman-Haham, I. et al. Structure of the radial spoke head and insights into its role in mechanoregulation of ciliary beating.Nat. Struct. Mol. Biol.28, 20–28(2021).

[33] [33] Chen, D. T. N., Heymann, M., Fraden, S., Nicastro, D. & Dogic, Z. ATP consumption of eukaryotic flagella measured at a single-cell level.Biophys. J.109, 2562–2573(2015).

[34] [34] Maekawa, M. et al. Stage-specific expression of mouse germ cell-less-1(mGCL-1), and multiple deformations during mgcl-1 deficient spermatogenesis leading to reduced fertility.Arch. Histol. Cytol.67, 335–347(2004).

[35] [35] Kimura, T. et al. Mouse germ cell-less as an essential component for nuclear integrity.Mol. Cell. Biol.23, 1304–1315(2003).

[36] [36] Mori, T. et al. CFAP47 is implicated in X-linked polycystic kidney disease.Kidney Int. Rep.9, 3580–3591(2024).

[37] [37] Zhang, X. et al. Differential requirements of IQUB for the assembly of radial spoke 1 and the motility of mouse cilia and flagella.Cell Rep.41, 111683(2022).

[38] [38] Sapiro, R. et al. Male infertility, impaired sperm motility, and hydrocephalus in mice deficient in sperm-associated antigen 6.Mol. Cell. Biol.22, 6298–6305(2002).

[39] [39] Miyata, H. et al. Testis-enriched kinesin KIF9 is important for progressive motility in mouse spermatozoa.FASEB J.34, 5389–5400(2020).

[40] [40] Tu, C. et al. Novel mutations in SPEF2 causing different defects between flagella and cilia bridge: the phenotypic link between MMAF and PCD.Hum. Genet.139, 257–271(2020).

[41] [41] Lechtreck, K. F. & Witman, G. B. Chlamydomonas reinhardtii hydin is a central pair protein required for flagellar motility.J. Cell Biol.176, 473–482(2007).

[42] [42] Lechtreck, K. F., Delmotte, P., Robinson, M. L., Sanderson, M. J. & Witman, G. B. Mutations in Hydin impair ciliary motility in mice.J. Cell Biol.180, 633–643(2008).

[43] [43] Wargo, M. J. & Smith, E. F. Asymmetry of the central apparatus defines the location of active microtubule sliding in Chlamydomonas flagella.Proc. Natl. Acad. Sci. USA100, 137–142(2003).

[44] [44] Zheng, S. et al. AreTomo: An integrated software package for automated markerfree, motion-corrected cryo-electron tomographic alignment and reconstruction.J. Struct. Biol. X6, 100068(2022).

[45] [45] Tegunov, D. & Cramer, P. Real-time cryo-electron microscopy data preprocessing with Warp.Nat. Methods16, 1146–1152(2019).

[46] [46] Tegunov, D., Xue, L., Dienemann, C., Cramer, P. & Mahamid, J. Multi-particle cryo-EM refinement with M visualizes ribosome-antibiotic complex at 3. 5 in cells.Nat. Methods18, 186–193(2021).

[47] [47] Lindemann, C. B. & Lesich, K. A. The mechanics of cilia and flagella: What we know and what we need to know.Cytoskeleton81, 648–668(2024).

[48] [48] Lindemann, C. B. & Lesich, K. A. The many modes of flagellar and ciliary beating: Insights from a physical analysis.Cytoskeleton78, 36–51(2021).

[49] [49] Lindemann, C. B., Orlando, A. & Kanous, K. S. The flagellar beat of rat sperm is organized by the interaction of two functionally distinct populations of dynein bridges with a stable central axonemal partition.J. Cell Sci.102, 249–260(1992).

[50] [50] Liao, H. Q. et al. WDR87 interacts with CFAP47 protein in the middle piece of spermatozoa flagella to participate in sperm tail assembly.Mol. Hum. Reprod.29, gaac042(2022).

[51] [51] Liu, M. et al. A novel mutation in CFAP47 causes male infertility due to multiple morphological abnormalities of the sperm flagella.Front. Endocrinol.14, 1155639(2023).

[52] [52] Ge, H. et al. Mutations in CFAP47, a previously reported MMAF causative gene, also contribute to the respiratory defects in patients with PCD.Mol. Genet. Genom. Med.12, e2278(2024).

[53] [53] Fontana, P. et al. Structure of cytoplasmic ring of nuclear pore complex by integrative cryo-EM and AlphaFold.Science376, eabm9326(2022).

[54] [54] Li, H. & Durbin, R. Fast and accurate short read alignment with Burrows-Wheeler transform.Bioinformatics25, 1754–1760(2009).

[55] [55] McKenna, A. et al. The Genome Analysis Toolkit: a MapReduce framework for analyzing next-generation DNA sequencing data.Genome Res.20, 1297–1303(2010).

[56] [56] Wang, K., Li, M. & Hakonarson, H. ANNOVAR: functional annotation of genetic variants from high-throughput sequencing data.Nucleic Acids Res.38, e164(2010).

[57] [57] Venkatraman, E. S. & Olshen, A. B. A faster circular binary segmentation algorithm for the analysis of array CGH data.Bioinformatics23, 657–663(2007).

[58] [58] Cooper, T. G. et al. World Health Organization reference values for human semen characteristics.Hum. Reprod. Update16, 231–245(2010).

[59] [59] World Health Organization.WHO Laboratory Manual for the Examination and Processing of Human Semen, 6th edition. (2021).

[60] [60] Gardner, D. K. & Lane, M. Culture and selection of viable blastocysts: a feasible proposition for human IVF?Hum. Reprod. Update3, 367–382(1997).

[61] [61] Alpha Scientists in Reproductive Medicine and ESHRE Special Interest Group of Embryology The Istanbul consensus workshop on embryo assessment: proceedings of an expert meeting.Hum. Reprod.26, 1270–1283(2011).

[62] [62] Hagen, W. J. H., Wan, W. & Briggs, J. A. G. Implementation of a cryo-electron tomography tilt-scheme optimized for high resolution subtomogram averaging.J. Struct. Biol.197, 191–198(2017).

[63] [63] Wu, C., Huang, X., Cheng, J., Zhu, D. & Zhang, X. High-quality, high-throughput cryo-electron microscopy data collection via beam tilt and astigmatism-free beam-image shift.J. Struct. Biol.208, 107396(2019).

[64] [64] Kremer, J. R., Mastronarde, D. N. & McIntosh, J. R. Computer visualization of three-dimensional image data using IMOD.J. Struct. Biol.116, 71–76(1996).

[65] [65] Castano-Diez, D., Kudryashev, M., Arheit, M. & Stahlberg, H. Dynamo: a flexible, user-friendly development tool for subtomogram averaging of cryo-EM data in high-performance computing environments.J. Struct. Biol.178, 139–151(2012).

[66] [66] Zivanov, J. et al. New tools for automated high-resolution cryo-EM structure determination in RELION-3.Elife7, e42166(2018).

[67] [67] He, J., Li, T. & Huang, S. Y. Improvement of cryo-EM maps by simultaneous local and non-local deep learning.Nat. Commun.14, 3217(2023).

[68] [68] Pettersen, E. F. et al. UCSF Chimera-a visualization system for exploratory research and analysis.J. Comput. Chem.25, 1605–1612(2004).

[69] [69] Pettersen, E. F. et al. UCSF ChimeraX: Structure visualization for researchers, educators, and developers.Protein Sci.30, 70–82(2021).

[70] [70] Jumper, J. et al. Highly accurate protein structure prediction with AlphaFold.Nature596, 583–589(2021).

[71] [71] Emsley, P., Lohkamp, B., Scott, W. G. & Cowtan, K. Features and development of Coot.Acta Crystallogr. D Biol. Crystallogr.66, 486–501(2010).

[72] [72] Liebschner, D. et al. Macromolecular structure determination using X-rays, neutrons and electrons: recent developments in Phenix.Acta Crystallogr. D Struct. Biol.75, 861–877(2019).

[73] [73] Dai, D., Ichikawa, M., Peri, K., Rebinsky, R. & Huy Bui, K. Identification and mapping of central pair proteins by proteomic analysis.Biophys. Physicobiol.17, 71–85(2020).