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.
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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.