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Homozygous mutations in SPEF2 induce multiple morphological abnormalities of the sperm flagella and male infertility
  1. Chunyu Liu1,2,3,4,
  2. Mingrong Lv4,5,6,
  3. Xiaojin He4,5,6,
  4. Yong Zhu1,
  5. Amir Amiri-Yekta7,8,9,
  6. Weiyu Li1,2,3,
  7. Huan Wu4,5,6,
  8. Zine-Eddine Kherraf7,8,
  9. Wangjie Liu1,2,3,
  10. Jingjing Zhang4,5,6,
  11. Qing Tan4,5,6,
  12. Shuyan Tang1,2,3,
  13. Yong-Jun Zhu10,
  14. Yading Zhong11,
  15. Caihua Li12,
  16. Shixiong Tian1,
  17. Zhiguo Zhang4,5,6,
  18. Li Jin1,
  19. Pierre Ray7,8,
  20. Feng Zhang1,2,3,4,
  21. Yunxia Cao4,5,6
  1. 1 Obstetrics and Gynecology Hospital, NHC Key Laboratory of Reproduction Regulation (Shanghai Institute of Planned Parenthood Research), State Key Laboratory of Genetic Engineering at School of Life Sciences, Fudan University, Shanghai, China
  2. 2 Shanghai Key Laboratory of Female Reproductive Endocrine Related Diseases, Shanghai, China
  3. 3 State Key Laboratory of Reproductive Medicine, Center for Global Health, School of Public Health, Nanjing Medical University, Nanjing, China
  4. 4 Reproductive Medicine Center, Department of Obstetrics and Gynecology, The First Affiliated Hospital of Anhui Medical University, Hefei, China
  5. 5 Anhui Province Key Laboratory of Reproductive Health and Genetics, Anhui Medical University, Hefei, China
  6. 6 Anhui Provincial Engineering Technology Research Center for Biopreservation and Artificial Organs, Hefei, China
  7. 7 Genetic Epigenetic and Therapies of Infertility, Institute for Advanced Biosciences, INSERM U1209, CNRS UMR 5309, Université Grenoble Alpes, Grenoble, France
  8. 8 Centre Hospitalier Universitaire de Grenoble, UM GI-DPI, Grenoble, France
  9. 9 Department of Genetics, Reproductive Biomedicine Research Center, Royan Institute for Reproductive Biomedicine, Academic Center for Education, Culture, and Research, Tehran, Iran
  10. 10 Department of Thoracic Surgery, Huashan Hospital, Fudan University, Shanghai, China
  11. 11 Department of Radiology, The First Affiliated Hospital of Anhui Medical University, Hefei, China
  12. 12 Genesky Biotechnologies Inc, Shanghai, China
  1. Correspondence to Professor Feng Zhang, Fudan University, Shanghai, China; zhangfeng{at}; Professor Yunxia Cao, Anhui Medical University, Hefei, China; caoyunxia6{at}


Background Male infertility due to multiple morphological abnormalities of the sperm flagella (MMAF) is a genetically heterogeneous disorder. Previous studies revealed several MMAF-associated genes, which account for approximately 60% of human MMAF cases. The pathogenic mechanisms of MMAF remain to be illuminated.

Methods and results We conducted genetic analyses using whole-exome sequencing in 50 Han Chinese probands with MMAF. Two homozygous stop-gain variants (c.910C>T (p.Arg304*) and c.3400delA (p.Ile1134Serfs*13)) of the SPEF2 (sperm flagellar 2) gene were identified in two unrelated consanguineous families. Consistently, an Iranian subject from another cohort also carried a homozygous SPEF2 stop-gain variant (c.3240delT (p.Phe1080Leufs*2)). All these variants affected the long SPEF2 transcripts that are expressed in the testis and encode the IFT20 (intraflagellar transport 20) binding domain, important for sperm tail development. Notably, previous animal studies reported spontaneous mutations of SPEF2 causing sperm tail defects in bulls and pigs. Our further functional studies using immunofluorescence assays showed the absence or a remarkably reduced staining of SPEF2 and of the MMAF-associated CFAP69 protein in the spermatozoa from SPEF2-affected subjects.

Conclusions We identified SPEF2 as a novel gene for human MMAF across the populations. Functional analyses suggested that the deficiency of SPEF2 in the mutated subjects could alter the localisation of other axonemal proteins.

  • spef2
  • sperm
  • infertility
  • exome
  • sequencing

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  • ChL, ML, XH, YoZ and PR contributed equally.

  • Contributors ChL, ML, XH, PR and FZ designed the study. ML, XH, AA-Y, HW, JZ, QT, Y-JZ, YaZ, ZZ and YC provided patients’ data and performed clinical assessments. ChL, YoZ, AA-Y, WeL, Z-EK, WaL and STi conducted the experiments. ChL, ML, XH, YoZ, HW, Z-EK, STa, CaL, LJ, PR, FZ and YC analysed the data. ChL, XH, PR, FZ and YC wrote the manuscript. PR, FZ and YC supervised the study.

  • Funding This work was supported by the National Natural Science Foundation of China (31625015, 31521003 and 81601340), Special Foundation for Development of Science and Technology of Anhui Province (2017070802D150 and YDZX20183400004194), Foundation of the Education Department of Anhui Province (KJ2016A370), Natural Science Foundation of Anhui Province (1708085QC59), Shanghai Medical Center of Key Programs for Female Reproductive Diseases (2017ZZ01016), and Shanghai Municipal Science and Technology Major Project (2017SHZDZX01).

  • Competing interests None declared.

  • Patient consent for publication All patients gave their signed informed consent for this study.

  • Ethics approval Declaration of Helsinki and approved by the Ethical Committees of the centers participating in this study.

  • Provenance and peer review Not commissioned; externally peer reviewed.