Biallelic CFAP61 variants cause male infertility in humans and mice with severe oligoasthenoteratozoospermia

Background The genetic causes for most male infertility due to severe oligoasthenoteratozoospermia (OAT) remain unclear. Objective To identify the genetic cause of male infertility characterised by OAT. Methods Variant screening was performed by whole-exome sequencing from 325 infertile patients with OAT and 392 fertile individuals. In silico and in vitro analyses were performed to evaluate the impacts of candidate disease-causing variants. A knockout mouse model was generated to confirm the candidate disease-causing gene, and intracytoplasmic sperm injection (ICSI) was used to evaluate the efficiency of clinical treatment. Results We identified biallelic CFAP61 variants (NM_015585.4: c.1654C>T (p.R552C) and c.2911G>A (p.D971N), c.144–2A>G and c.1666G>A (p.G556R)) in two (0.62%) of the 325 OAT-affected men. In silico bioinformatics analysis predicted that all four variants were deleterious, and in vitro functional analysis confirmed the deleterious effects of the mutants. Notably, H&E staining and electron microscopy analyses of the spermatozoa revealed multiple morphological abnormalities of sperm flagella, the absence of central pair microtubules and mitochondrial sheath malformation in sperm flagella from man with CFAP61 variants. Further immunofluorescence assays revealed markedly reduced CFAP61 staining in the sperm flagella. In addition, Cfap61 -deficient mice showed the OAT phenotype, suggesting that loss of function of CFAP61 was the cause of OAT. Two individuals accepted ICSI therapy using their own ejaculated sperm, and one of them succeeded in fathering a healthy baby. Conclusions Our findings indicate that CFAP61 is essential for spermatogenesis and that biallelic CFAP61 variants lead to male infertility in humans and mice with OAT.