Bi-allelic variants in chromatoid body protein TDRD6 cause spermiogenesis defects and severe oligoasthenoteratozoospermia in humans.

Background: The association between the variants and human infertility remains unclear, as only one homozygous missense variant of was found to be associated with oligoasthenoteratozoospermia (OAT).

Methods: Whole-exome sequencing and Sanger sequencing were employed to identify potential pathogenic variants of in infertile men. Histology, immunofluorescence, immunoblotting and ultrastructural analyses were conducted to clarify the structural and functional abnormalities of sperm in mutated patients. -knockout mice were generated using the CRISPR-Cas9 system. Total RNA-seq and single-cell RNA-seq (scRNA-seq) analyses were used to elucidate the underlying molecular mechanisms, followed by validation through quantitative RT-PCR and immunostaining. Intracytoplasmic sperm injection (ICSI) was also used to assess the efficacy of clinical treatment.

Results: Bi-allelic variants were identified in five unrelated Chinese individuals with OAT, including homozygous loss-of-function variants in two consanguineous families. Notably, besides reduced concentrations and impaired motility, a significant occurrence of acrosomal hypoplasia was detected in multiple spermatozoa among five patients. Using the -deficient mice, we further elucidate the pivotal role of TDRD6 in spermiogenesis and acrosome identified. In addition, the mislocalisation of crucial chromatoid body components DDX4 (MVH) and UPF1 was also observed in round spermatids from patients harbouring variants. ScRNA-seq analysis of germ cells from a patient with variants revealed that TDRD6 regulates mRNA metabolism processes involved in spermatid differentiation and cytoplasmic translation.

Conclusion: Our findings strongly suggest that TDRD6 plays a conserved role in spermiogenesis and confirms the causal relationship between variants and human OAT. Additionally, this study highlights the unfavourable ICSI outcomes in individuals with bi-allelic variants, providing insights for potential clinical treatment strategies.