Department of Biomedical Sciences, College of Veterinary Medicine, Cornell University, Ithaca, New York 14853, USA.
Biol Reprod. 2012 Oct 18;87(4):88. doi: 10.1095/biolreprod.112.099721. Print 2012 Oct.
Three Binder of SPerm proteins (BSP1, BSP3, BSP5) are secreted by bovine seminal vesicles into seminal plasma and adsorbed onto sperm. When sperm inseminated into the female reach the oviduct, the BSP proteins bind them to its epithelial lining, forming a sperm storage reservoir. Previously, we reported that binding of capacitated sperm to oviductal epithelium in vitro is lower than that of uncapacitated sperm and we proposed that reduced binding was due to loss of BSP proteins during capacitation. Because of differences in amino acid sequences, we predicted that each BSP would respond differently to capacitating conditions. To test whether all three BSP proteins were lost from sperm during capacitation and whether the kinetics of loss differed among the three BSP proteins, ejaculated bull sperm were incubated under various capacitating conditions, and then the amounts of BSP proteins remaining on the sperm were assayed by Western blotting. Capacitation was assayed by analysis of protein tyrosine phosphorylation. While loss of BSP1 was not detected, most of the BSP5 was lost from sperm during incubation in TALP medium, even without addition of the capacitation enhancers heparin and dbcAMP-IBMX. Surprisingly, a smaller molecular mass was detected by anti-BSP3 antibodies in extracts of incubated sperm. Its identity was confirmed as BSP3 by mass spectrometry, indicating that BSP3 undergoes modification on the sperm surface. These changes in the composition of BSP proteins on sperm could play a role in releasing sperm from the storage reservoir by modifying sperm interactions with the oviductal epithelium.
三种 Binder of SPerm 蛋白（BSP1、BSP3、BSP5）由牛精囊分泌到精液中，并吸附在精子上。当精子在雌性生殖道中受精时，BSP 蛋白将其结合到上皮细胞衬里上，形成精子储存库。此前，我们报道体外受精的精子与输卵管上皮的结合能力低于未受精的精子，我们提出这种结合能力降低是由于在受精过程中 BSP 蛋白的丢失。由于氨基酸序列的差异，我们预测每个 BSP 蛋白对受精条件的反应都不同。为了测试受精过程中所有三种 BSP 蛋白是否从精子中丢失，以及丢失的动力学是否在三种 BSP 蛋白之间存在差异，我们将射出的公牛精子在不同的受精条件下孵育，然后通过 Western blot 测定剩余的 BSP 蛋白量。通过分析蛋白酪氨酸磷酸化来检测受精。虽然没有检测到 BSP1 的丢失，但在 TALP 培养基中孵育时，大部分 BSP5 从精子中丢失，即使没有添加肝素和 dbcAMP-IBMX 等受精增强剂。令人惊讶的是，在孵育精子的提取物中，抗 BSP3 抗体检测到较小分子量的物质。通过质谱法确定其为 BSP3，表明 BSP3 在精子表面发生修饰。这些精子上 BSP 蛋白组成的变化可能通过改变精子与输卵管上皮的相互作用，在从储存库中释放精子方面发挥作用。
