SPACA4 regulates the structure and molecular basis of spermatid maturation and ultimately affects sperm quality in mice

Sperm quality is crucial for sperm function and can even affect embryo quality and offspring health. Spermatid maturation is extremely complex, as spermatids undergo morphological changes, laying the foundation for the execution of sperm function. The function of sperm acrosome-associated 4 (SPACA4) in spermatogenesis is not well known. The present study revealed that SPACA4 was specifically expressed in the acrosomes and cytoplasm of mouse spermatids. Spaca4 knockout mice demonstrated that the loss of SPACA4 led to male subfertility. The quality of mature sperm was abnormal in Spaca4-/- mice, manifested by decreased motility and multiple deformities. Spaca4-/- sperm exhibited irregular nuclear shapes, abnormal nuclei with vacuoles, missing or incompletely fused acrosomes, and multiple cross-sections enclosed in the same sperm cell membrane. Electron microscopy and molecular expression analyses of testicles revealed that the loss of SPACA4 affected the differentiation of the acrosome, acroplaxome, and manchette, resulting in abnormalities in nuclear elongation, chromatin condensation, and flagellar development. Interestingly, SPACA4 did not regulate spermiogenesis via the acetylcholine signaling pathway. Analysis of the differential protein expression profile revealed that the expression of 9 proteins was significantly decreased in Spaca4-/- spermatids. A decreased protein, transformation-related protein 53 target 5 (TRP53TG5), was knocked down in spermatids and found that the phenotype was consistent with Spaca4 knockout mice. These results revealed that the absence of SPACA4 leads to abnormal spermatid maturation and affects sperm quality in mice. Abnormal sperm quality in Spaca4-/- mice results in decreased sperm capacitation and a decreased acrosome response, ultimately affecting the fertility of male mice.