Abou-Haïla A, Fouquet J P, Tulsiani D R
Laboratoire de Biologie Cellulaire, Université René Descartes, 45, rue des Saints-Pères, Paris Cedex 06, 75270, France.
Exp Cell Res. 1999 Feb 25;247(1):48-60. doi: 10.1006/excr.1998.4355.
Spermatozoa released from the seminiferous tubules are terminally differentiated cells with no known synthetic activity. Their components are synthesized in the spermatogenic cells during spermatogenesis. In this study, we report the characterization and immunolocalization of beta-glucuronidase in mouse testicular germ cells and spermatozoa. The enzyme is an exoglycohydrolase with dual localization, being present in lysosomes and endoplasmic reticulum of several mouse and rat tissues. The purified germ cell preparations (spermatocytes, round spermatids, and condensed/elongated spermatids) when assayed for beta-glucuronidase activity showed that the spermatocytes contained five times more enzyme activity per cell than the spermatids. Polyacrylamide gel electrophoresis, carried out under native and denaturing conditions, demonstrated that the germ cells express only the lysosomal form of the enzyme (pI 5.5-6.0) with a subunit molecular mass of 74 kDa. Immunocytochemical studies revealed a positive reaction in the Golgi membranes, Golgi-associated vesicles, and lysosomes of late spermatocytes (pachytene spermatocytes) and a stage-specific localization during spermiogenesis. The forming or formed acrosome of the elongated spermatids (stages 9-16) and epididymal spermatozoa was highly immunopositive. Comparison of immunoprecipitation curves and kinetic properties of the enzyme present in spermatocytes and spermatozoa revealed no major differences. Taken together, our results demonstrate that beta-glucuronidase activities present in the lysosomes of spermatocytes and the sperm acrosome are kinetically and immunologically similar.
从生精小管释放的精子是终末分化细胞，没有已知的合成活性。它们的成分在精子发生过程中由生精细胞合成。在本研究中，我们报告了小鼠睾丸生殖细胞和精子中β-葡萄糖醛酸酶的特性和免疫定位。该酶是一种具有双重定位的外切糖苷酶，存在于几种小鼠和大鼠组织的溶酶体和内质网中。对纯化的生殖细胞制剂（精母细胞、圆形精子细胞和浓缩/伸长精子细胞）进行β-葡萄糖醛酸酶活性检测，结果显示精母细胞每个细胞的酶活性比精子细胞高五倍。在天然和变性条件下进行的聚丙烯酰胺凝胶电泳表明，生殖细胞仅表达该酶的溶酶体形式（pI 5.5 - 6.0），亚基分子量为74 kDa。免疫细胞化学研究显示，晚期精母细胞（粗线期精母细胞）的高尔基体膜、高尔基体相关小泡和溶酶体呈阳性反应，并且在精子形成过程中具有阶段特异性定位。伸长精子细胞（9 - 16期）和附睾精子的正在形成或已形成的顶体高度免疫阳性。对精母细胞和精子中该酶的免疫沉淀曲线和动力学特性进行比较，未发现重大差异。综上所述，我们的结果表明，精母细胞溶酶体和精子顶体中存在的β-葡萄糖醛酸酶活性在动力学和免疫学上相似。
