Sperm Function in Fertilization Events

描述(申请人提供)：附睾上皮产生特定区域的管腔环境，促进睾丸后精子运动和受精能力的发展，并在附睾尾储存期间保持精子活力。在识别附睾对精子成熟的贡献方面已经取得了实质性的进展，然而它在储存过程中保持精子活力的机制还不是很清楚。我们已经鉴定出仓鼠尾部附睾主细胞的一种潜在的独特的分泌蛋白，它与有缺陷的管腔精子和精子片段特异性结合并聚合成一个围绕着有缺陷的管腔精子和精子片段的死亡茧，从而将它们从可存活的精子群中分离出来。这种蛋白质被称为eFGL(附睾纤维蛋白原样蛋白)，是一个由三个亚基组成的二硫键连接的寡聚体。两个eFGL亚基已经被测序，并被鉴定为纤维蛋白原相关蛋白：FGL2(纤维蛋白原样蛋白-2)是一个65 kDa的丝氨酸蛋白酶，FG11(纤维蛋白原样蛋白-1)是一个功能未知的33 kDa多肽。第三个亚基dc45(死亡茧45 kDa多肽)尚未鉴定。我们推测，eFGL代表了一种新的保护机制，它特异性地识别和掩盖有缺陷的精子，防止他们释放水解酶或抗原，如果他们逃脱血-附睾屏障，可能会对可存活的精子群体产生有害影响，或者启动炎症或免疫反应。本研究的具体目的是：1.确定eFGL是否仅由附睾主细胞表达和分泌。2：确定缺陷精子中哪些细胞器与附睾FGL寡聚体结合。3：确定eFGL与特定精子配体结合是否促进精子死亡和/或死亡茧聚合。4：通过靶向干扰小鼠的fgl2基因来确定eFGL的功能。

DESCRIPTION (provided by applicant): The epididymal epithelium produces a region-specific luminal environment that promotes the post-testicular development of sperm motility and fertilizing capacity and maintains sperm viability during storage in the cauda epididymidis. Substantial progress has been made on identifying epididymal contributions to sperm maturation, however the mechanisms by which it maintains sperm viability during storage is less well understood. We have identified a potentially unique secretory protein of hamster cauda epididymal principal cells which specifically binds to, and polymerizes into a "death cocoon" surrounding defective luminal spermatozoa and sperm fragments, thereby segregating them from the viable sperm population. This protein termed eFGL (for epididymal fibrinogen-like protein) is a disulfide-linked oligomer of three subunits. Two eFGL subunits have been sequenced and identified as fibrinogen-related proteins: fgl2 (fibrinogen-like protein-2) a 65 kDa serine-protease and fgll (fibrinogen-like protein-1) a 33 kDa polypeptide of unknown function. The third subunit dc45 (for death cocoon 45 kDa polypeptide) has not been identified. We hypothesize that eFGL represents a novel protective mechanism which specifically recognizes and masks defective spermatozoa preventing their release ofhydrolases or antigens which could exert deleterious effects on the viable sperm population or initiate inflammatory or immune responses if they escape the blood-epididymis barrier. The specific aims of this study are: 1: To determine whether eFGL is expressed and secreted by only epididymal principal cells. 2: To determine which organelles of defective spermatozoa bind the epididymal FGL oligomer. 3: To determine if eFGL binding to specific sperm ligands promotes sperm death and/or death cocoon polymerization.4: To define the function of eFGL by targeted disruption of the fgl2 gene in mice.