Biallelic variants in IQCN cause sperm flagellar assembly defects and male infertility

• Published in: Human reproduction (Oxford) 2023-07, Vol.38 (7), p.1390-1398
• Main Authors: Li, Qi, Wang, Yize, Zheng, Wei, Guo, Jing, Zhang, Shunji, Gong, Fei, Lu, Guang-Xiu, Lin, Ge, Dai, Jing
• Format: Article
• Language: English
• Subjects: Animals; Humans; Infertility, Male - genetics; Infertility, Male - metabolism; Male; Mice; Semen - metabolism; Sperm Motility - genetics; Sperm Tail - metabolism; Spermatozoa - metabolism; Spermatozoa - pathology
• ISSN: 0268-1161; 1460-2350
• DOI: 10.1093/humrep/dead079

Abstract STUDY QUESTION What is the effect of defects in the manchette protein IQ motif-containing N (IQCN) on sperm flagellar assembly? SUMMARY ANSWER Deficiency in IQCN causes sperm flagellar assembly defects and male infertility. WHAT IS KNOWN ALREADY The manchette is a transient structure that is involved in the shaping of the human spermatid nucleus and protein transport within flagella. Our group recently reported that the manchette protein IQCN is essential for fertilization. Variants in IQCN lead to total fertilization failure and defective acrosome structure phenotypes. However, the function of IQCN in sperm flagellar assembly is still unknown. STUDY DESIGN, SIZE, DURATION Fifty men with infertility were recruited from a university-affiliated center from January 2014 to October 2022. PARTICIPANTS/MATERIALS, SETTING, METHODS Genomic DNA was extracted from the peripheral blood samples of all 50 individuals for whole-exome sequencing. The ultrastructure of the spermatozoa was assessed by transmission electron microscopy. Computer-assisted sperm analysis (CASA) was used to test the parameters of curvilinear velocity (VCL), straight-line velocity (VSL), and average path velocity (VAP). An Iqcn knockout (Iqcn−/−) mouse model was generated by CRISPR–Cas9 technology to evaluate sperm motility and the ultrastructure of the flagellum. Hyperactivation and sperm fertilizing ability were assessed in a mouse model. Immunoprecipitation followed by liquid chromatography–mass spectrometry was used to detect IQCN-binding proteins. Immunofluorescence was used to validate the localization of IQCN-binding proteins. MAIN RESULTS AND THE ROLE OF CHANCE Biallelic variants in IQCN (c.3913A>T and c.3040A>G; c.2453_2454del) were identified in our cohort of infertile men. The sperm from the affected individuals showed an irregular ‘9 + 2’ structure of the flagellum, which resulted in abnormal CASA parameters. Similar phenotypes were observed in Iqcn−/− male mice. VSL, VCL, and VAP in the sperm of Iqcn−/− male mice were significantly lower than those in Iqcn+/+ male mice. Partial peripheral doublet microtubules (DMTs) and outer dense fibers (ODFs) were absent, or a chaotic arrangement of DMTs was observed in the principal piece and end piece of the sperm flagellum. Hyperactivation and IVF ability were impaired in Iqcn−/− male mice. In addition, we investigated the causes of motility defects and identified IQCN-binding proteins including CDC42 and the intraflagellar transport