Population-based genetic analysis in infertile men reveals novel mutations of ADAD family members in patients with impaired spermatogenesis.

The testis-specific adenosine deaminase domain-containing (ADAD) protein family, including ADAD1 and ADAD2, has been confirmed to be essential in mouse male fertility. However, the roles of ADAD1 and ADAD2 in human reproductive biology are unclear. Herein, whole-exome sequencing was conducted for 337 infertile patients to detect pathogenic variants in ADAD1 and ADAD2. Importantly, a novel deleterious biallelic variant of NM_001159285.2:c.1408G > T (p.V470F) and NM_001159285.2:c.1418A > G (p.E473G) in ADAD1 and a pathogenic homozygous missense variant of NM_001145400.2:c.1381C > T (p.R461W) in ADAD2 were identified in this infertile cohort with frequencies of 0.29 (1/337) and 0.59% (2/337), respectively. Electron microscopy revealed an abnormal morphology and severely disorganized ultrastructure of sperm from the patients. Immunofluorescence and western blotting showed a sharp decrease in ADAD1 and ADAD2 expression in sperm from the patients. Mechanistically, bioinformatics analysis suggested that ADAD2 interacts with DNAH17. Furthermore, we demonstrated that the expression of DNAH17 was markedly downregulated in the sperm of patients harboring ADAD2 variants. In addition, the expression of several autophagy regulators was significantly disrupted in the sperm of patients harboring ADAD2 variants. In conclusion, we identified novel ADAD1 and ADAD2 variants in three infertile patients from a large infertile cohort, first providing evidence that ADAD1 and ADAD2 variants might be a candidate genetic cause of human male infertility. Moreover, an important new dimension to our understanding of the genotype-phenotype correlations between the ADAD gene family and male infertility in humans has been uncovered, providing valuable information for the genetic diagnosis of male infertility.