Miranda-Vizuete Antonio, Tsang Katie, Yu Yang, Jiménez Alberto, Pelto-Huikko Markku, Flickinger Charles J, Sutovsky Peter, Oko Richard
Center for Biotechnology, Department of Biosciences at Novum, Karolinska Institutet, S-14157 Huddinge, Sweden.
J Biol Chem. 2003 Nov 7;278(45):44874-85. doi: 10.1074/jbc.M305475200. Epub 2003 Aug 7.
Thioredoxins compose a growing family of proteins that participate in different cellular processes via redox-mediated reactions. We report here the cloning, developmental expression, and location of murid Sptrx-2. Mouse and rat SPTRX-2 proteins display a high homology to their human ortholog in the thioredoxin and NDP kinase domains, and the coding genes are located at syntenic positions. Northern blotting and in situ hybridization confirmed the testis-specific expression of murine Sptrx-2 mRNA, mostly in round spermatids. Immunohistochemical analysis of the 19 steps of rat spermiogenesis showed that SPTRX-2 expression becomes prominent in the cytoplasmic lobe of step 15-18 spermatids and diminishes in step 19 just before spermiation. However, in the spermatid tail, SPTRX-2 immunoreactivity increased from step 15 to 19 and was confined to the principal piece. By immunogold electron microscopy, SPTRX-2 was first detected scattered throughout the cytoplasm of the axoneme in step 14-15 spermatids, but began to be incorporated by step 16 into the fibrous sheath (FS). During steps 17-18, the labeling increased over the ribs and columns of the assembled FS. It peaked in step 19 and remained in the FS of epididymal spermatozoa. Immunoblots of isolated FS obtained from spermatozoa confirmed that SPTRX-2 is an integral component of the FS and a post-obstruction autoantigen in vasectomized rats. Our data indicate that SPTRX-2 incorporation into the FS lags well behind FS assembly, suggesting it is required during the final stages of sperm tail maturation in the testis and/or epididymis, where extensive disulfide bonding of FS proteins occurs.
硫氧还蛋白构成了一个不断扩大的蛋白质家族，它们通过氧化还原介导的反应参与不同的细胞过程。我们在此报告鼠源Sptrx-2的克隆、发育表达及定位。小鼠和大鼠的SPTRX-2蛋白在硫氧还蛋白和NDP激酶结构域与其人类直系同源物具有高度同源性，且编码基因位于同线位置。Northern印迹和原位杂交证实了小鼠Sptrx-2 mRNA在睾丸中的特异性表达，主要在圆形精子细胞中表达。对大鼠精子发生19个阶段的免疫组织化学分析表明，SPTRX-2的表达在第15 - 18阶段精子细胞的细胞质叶中变得显著，并在第19阶段精子释放前减少。然而，在精子尾部，SPTRX-2免疫反应性从第15阶段到第19阶段增加，并局限于主段。通过免疫金电子显微镜观察，在第14 - 15阶段精子细胞中，首先在轴丝的整个细胞质中散在检测到SPTRX-2，但在第16阶段开始被整合到纤维鞘（FS）中。在第17 - 18阶段，在组装好的FS的肋和柱上标记增加。在第19阶段达到峰值，并保留在附睾精子的FS中。从精子中分离得到的FS的免疫印迹证实，SPTRX-2是FS的一个组成成分，也是输精管结扎大鼠梗阻后自身抗原。我们的数据表明，SPTRX-2整合到FS的过程远远落后于FS的组装，这表明它在睾丸和/或附睾中精子尾部成熟的最后阶段是必需的，在这个阶段FS蛋白会发生广泛的二硫键结合。
