Identification and Validation of Yak (Bos grunniens) Frozen–Thawed Sperm Proteins Associated with Capacitation and the Acrosome Reaction

To achieve fertilization, mammalian spermatozoa must undergo capacitation and the acrosome reaction (AR) within the female reproductive tract. However, the effects of cryopreservation on sperm maturation and fertilizing potential have yet to be established. To gain insight into changes in protein levels within sperm cells prepared for use in the context of fertilization, a comprehensive quantitative proteomic profiling approach was used to analyze frozen–thawed Ashidan yak spermatozoa under three sequential conditions: density gradient centrifugation-based purification, incubation in a capacitation medium, and treatment with the calcium ionophore A23187 to facilitate AR induction. In total, 3280 proteins were detected in these yak sperm samples, of which 3074 were quantified, with 68 and 32 being significantly altered following sperm capacitation and AR induction. Differentially abundant capacitation-related proteins were enriched in the metabolism and PPAR signaling pathways, while differentially abundant AR-related proteins were enriched in the AMPK signaling pathway. These data confirmed a role for superoxide dismutase 1 (SOD1) as a regulator of sperm capacitation while also offering indirect evidence that heat shock protein 90 alpha (HSP90AA1) regulates the AR. Together, these findings offer a means whereby sperm fertility-related marker proteins can be effectively identified. Data are available via Proteome Xchange with identifier PXD035038.