"Sperm maturation and capacitation: Roles of Chaperone Proteins, Clusterin and Hsp70"

提高受精效率将导致农业动物生产的更大成功。精子是在睾丸中产生的，但睾丸精子不能使卵子受精。它们在通过附睾(称为精子成熟)和在雌性生殖道内迁移(称为获能)的过程中必须进行生化修饰。附睾上皮细胞分泌的大量卵细胞外基质透明带(ZP)结合蛋白沉积在中转精子表面。其中一些蛋白质倾向于聚集，在很小的附睾体体积内，它们可能在没有救援机制的情况下沉淀。聚集素(CLU)也由附睾上皮细胞分泌，具有伴侣性质，并倾向于在疏水(非极性，防水)结构域与其他蛋白质结合。我们认为，CLU可以阻止ZP结合蛋白的沉淀，并伴随它们到达精子表面。将进行生化实验，以验证CLU可以防止ZP结合蛋白(如芳基硫酸酯酶A，SED1)的沉淀/聚集，并将通过比较CLU基因缺失小鼠和野生型小鼠精子上ZP结合蛋白的存在来评估CLU的伴侣活性。在获能过程中，我们最近的结果表明，ZP结合蛋白以复合体的形式聚集在一起，另一个伴侣Hsp70蛋白可能在这种聚集中发挥作用。我们将通过质谱学鉴定复合体中的ZP结合蛋白，并通过评估未获能精子经Hsp70处理后是否具有更强的受精能力(获能)来验证Hsp70的作用。我们建议的工作结果可以转化为提高某些ZP结合蛋白水平较低的精子的受精能力。他们将被CLU+所需的ZP结合蛋白以及Hsp70处理，以促进复合体的形成。我们的研究还将对精子成熟和获能过程有更好的了解。

Raising efficiency in fertilization would lead to increased success in agricultural animal production. Sperm are produced in the testis, but testicular sperm cannot fertilize an egg. They have to undergo biochemical modifications during their transit through the epididymis (called sperm maturation) and their migration inside the female reproductive tract (called capacitation). A number of egg extracellular matrix, zona pellucida (ZP), binding proteins secreted by epididymal epithelial cells deposit onto the transit sperm surface. Some of these proteins tend to aggregate and within a small epididymal volume, they may precipitate without rescuing mechanisms. Clusterin (CLU), also secreted by epididymal epithelial cells, has a chaperone property and propensity to bind to other proteins at their hydrophobic (non-polar, water repellant) domains. We propose that CLU prevents ZP binding proteins from precipitation and chaperones them to the sperm surface. Biochemical experiments will be performed to validate that CLU can prevent precipitation/aggregation of ZP binding proteins (e.g., arylsulfatase A, SED1), and CLU's chaperone activity will be assessed by comparing the presence of ZP binding proteins on sperm in Clu genetically null mice versus that in wild type mice. During capacitation, our recent results suggest that ZP binding proteins conglomerate as complexes, and another chaperone Hsp70 protein may play roles in this conglomeration. We will identify ZP binding proteins in the complex by mass spectrometry and validate the role of Hsp70 by assessing whether sperm have more fertilizing ability (capacitated) after treating non-capacitated sperm with Hsp70. Results from our proposed work can be translated to enhance fertilizing ability of sperm that have low levels of certain ZP binding proteins. They will be treated with CLU + the desired ZP binding proteins, as well as Hsp70 to enhance the complex formation. Our study will also give a better understanding of sperm maturation and capacitation processes.