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.

为实现受精过程，哺乳动物精子必须在雌性生殖道内完成获能（capacitation）与顶体反应（acrosome reaction, AR）。然而，冷冻保存对精子成熟及受精潜能的影响尚未阐明。为深入解析受精相关制备流程中精子细胞内蛋白质水平的变化规律，本研究采用全面定量蛋白质组学分析策略，对经过三个连续处理阶段的冻融阿什丹牦牛精子开展分析：基于密度梯度离心的纯化步骤、获能培养基孵育，以及使用钙离子载体A23187诱导顶体反应。本研究共在该批牦牛精子样本中检测到3280种蛋白质，其中3074种可被定量；在精子获能与顶体反应诱导后，分别有68种和32种蛋白质发生显著丰度变化。获能相关差异丰度蛋白质富集于代谢通路与过氧化物酶体增殖物激活受体（PPAR）信号通路，而顶体反应相关差异丰度蛋白质则富集于AMPK信号通路。本研究数据证实超氧化物歧化酶1（superoxide dismutase 1, SOD1）可作为精子获能的调控因子，同时间接证明热休克蛋白90α（heat shock protein 90 alpha, HSP90AA1）可调控顶体反应。综上，本研究结果可为有效鉴定精子受精能力相关标记蛋白质提供可行途径。相关数据集已通过Proteome Xchange公开，标识符为PXD035038。