Molecular Mechanism of Functional Regulation of Spermatiogenic Cells by Interaction with Other Somatic Cells and Eggs

为了阐明其他体细胞和卵子对小鼠雄性配子的功能调控，我们研究了小鼠精子发生、精子成熟、精子从子宫到输卵管转运和受精过程中的细胞相互作用。实验结果如下：(1)从小鼠睾丸cDNA文库中鉴定出了睾丸特异性多聚(a)聚合酶（TPAP）的cDNA克隆，TPAP是一种负责生精细胞中已有mrna的细胞质聚腺苷化的候选分子。缺乏TPAP的雄性小鼠导致精子发生阻滞和生精细胞凋亡增加。单倍体圆形精子发育阶段前细胞凋亡的增加可能不是由于生精阻滞的直接影响，而是由于光信号通过Sertoli细胞转导到生精细胞的早期阶段。(2)在小鼠中，两种不同的ADAM1亚型。ADAM1a和ADAM1b是在睾丸中产生的。ADAM1a局限于睾丸生殖细胞的内质网，而附睾精子仅在质膜上含有ADAM1b。小鼠ADAM1a缺失导致雄性不育，因为精子从子宫通过子宫-输卵管连接处迁移到输卵管的能力严重受损。然而，adam1a缺失小鼠的附睾精子能够在体外与卵积云完整、透明带完整的卵子受精，尽管卵积云细胞的分散延迟，与透明带的粘附/结合减少。这些结果表明，ADAM1a/ADAM2受精蛋白复合物可能调节包括ADAM3在内的特定精子蛋白的选择性运输，这些蛋白参与了从睾丸生殖细胞内质网向输卵管的迁移、卵泡细胞的分散以及与透明带的结合，这些蛋白从睾丸生殖细胞的内质网到细胞表面。(3)缺乏精子PH-20的雄性小鼠仍然具有生育能力，尽管从积云团中分散积云细胞的能力降低。精子提取物的Western blot分析显示，除PH-20外，其他透明质酸酶(s)可能存在于精子顶体中。这些数据提供了至少在小鼠中PH-20不是受精所必需的证据，并表明其他透明质酸酶可能在精子穿透积云细胞层和/或卵细胞透明带中发挥重要作用，可能与PH-20合作。少

To elucidate the functional regulation of mouse male gamete by other somatic cells and eggs, we have examined the cell interaction in spermatogenesis, sperm maturation, sperm transit from uterus to oviduct, and fertilization in mouse. The following experimental results have been obtained(1)cDNA clones encoding a testis-specific poly(A) polymerase, TPAP, a candidate molecule responsible for cytoplasmic polyadenylation of pre-existing mRNAs in spermatogenic cells have been identified from a mouse testis cDNA library. Male mice lacking TPAP resulted in the arrest of spermatogenesis and in the increase of apoptosis in spermatogenic cells. The increased apoptosis before the stage of haploid round spermatids may be due not to direct effects of the spermatogenic arrest, but to a transduction of the spoptotic signals to the earlier stages of spermatogenic cells through Sertoli cells.(2)In mouse, two different isoforms of ADAM1. ADAM1a and ADAM1b, are produced in the testis. ADAM1a is restricte … More dly localized within the endoplasmic reticulum of testicular germ cells, whereas epididymal sperm contain only ADAM1b on the plasma membrane. The loss of mouse ADAM1a resulted in the male infertility because of the severely impaired ability of sperm to migrate from the uterus into oviduct through the uterotubal junction. However, epididymal sperm of ADAM1a-deficient mice were capable of fertilizing the cumulus-intact, zona pellucida-intact eggs in vitro, despite the delayed dispersal of cumulus cells and the reduced adhesion/binding to the zona pellucida. These results suggest that the ADAM1a/ADAM2 fertilin complex may regulate the selective transport of specific sperm proteins including ADAM3, which are involved in the migration into the oviduct, in the dispersal of cumulus cells, and in the binding to the zona pellucida, from the endoplasmic reticulum of testicular germ cells onto the cell surface.(3)Male mice lacking sperm PH-20 were still fertile, in spite of a reduced ability to disperse cumulus cells from the cumulus mass. Western blot analysis of sperm extracts revealed the presence of other hyaluronidase(s), except PH-20, presumably within the sperm acrosome. These data provide evidence that PH-20 is not essential for fertilization at least in the mouse, and suggest that the other hyaluronidase(s) may play an important role in the sperm penetration through the cumulus cell layer and/or the egg zona pellucida, possibly in cooperation with PH-20. Less