ELAPOR1 regulates VPS54-mediated GARP complex formation and proacrosomal vesicle fusion during spermatogenesis.

RATIONALE: Acrosomal developmental defects are associated with severe sperm morphogenetic abnormalities and male infertility; however, the specific molecular determinants underlying these defects remain poorly defined. Endosome/lysosome-associated apoptosis and autophagy regulator 1 (ELAPOR1) is important for cellular membrane dynamics and organelle functions, indicating that it plays an essential role in spermatogenesis, which needs further investigation. This study aimed to explore the specific role of ELAPOR1 in spermatogenesis and male fertility. METHODS: Single-cell RNA-sequencing datasets from human testes were obtained to investigate the expression of ELAPOR1 in specific cell types. An Elapor1 germ cell-specific knockout mouse line (Elapor1(cKO) ) was established, and the morphology and function of the testis and sperm were assessed through breeding capacity, fertilization capacity, testicular histology, sperm staining, concentration and motility, immunofluorescence, quantitative PCR, immunoblotting, and transmission electron microscopy analyses. Moreover, mass spectrometry and enrichment analyses were employed to identify the proteins that interact with ELAPOR1. The interactions were verified through proximity labeling, coimmunoprecipitation, and immunofluorescence staining. RESULTS: ELAPOR1 is highly expressed and its protein colocalized with the acrosome during the early stage of acrosome formation. Elapor1(cKO) mice produced deformed sperm with decreased concentration, impaired motility, and defective fertilization capacity. Moreover, ELAPOR1 deficiency led to impaired fusion of proacrosomal vesicles. Mechanistically, ELAPOR1 functioned in regulating the transport of the Golgi and early endosome-related vesicles. It interacted with VPS54 and affected VPS54-associated assembly of the GARP complex in the testis. CONCLUSIONS: Our findings reveal the essential role of ELAPOR1 in acrosome formation during spermatogenesis and male fertility. ELAPOR1 potentially influences the trafficking, integration, and fusion of proacrosomal vesicles through VPS54-mediated GARP complex assembly. These findings provide novel insights into the interaction of the ELAPOR1-GARP complex in acrosome-related reproductive failure, suggesting that Elapor1 deficiency or mutation could be considered a potential genetic risk factor for human infertility.