Germ cell-Sertoli cell interactions: analysis of the biosynthesis and secretion of cyclic protein-2.
Germ cell-Sertoli cell interactions: analysis of the biosynthesis and secretion of cyclic protein-2.
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生殖细胞-支持细胞相互作用：循环蛋白 2 的生物合成和分泌分析。
DOI：
10.1016/0012-1606(88)90412-5
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发表时间：
1988
影响因子：
2.7
通讯作者：
Wright,WW
中科院分区：
文献类型：
--
作者：
Wright,WW
在大鼠完整的VI期和VII期曲细精管中，成熟的Sertoli细胞在体外分泌大量的环蛋白-2（CP-2）。这种阶段依赖性分泌使我们假设这种分子的生物合成受到特定发育状态的生殖细胞的刺激。为了探索这一假说，并检查CP-2的生物合成的步骤，我们产生了针对这种蛋白质的多克隆抗血清，并用它来分析CP-2的生物合成和分泌。对CP-2生物合成步骤的分析表明，其多肽核心代表了CP-2 mRNA的大部分（如果不是全部的话）翻译产物，并且单个精氨酸连接的寡糖与该核心连接。还进行了生精上皮周期特定阶段CP-2生物合成速率的分析。在存在[35 S]甲硫氨酸和从小管免疫沉淀的放射性标记CP-2的情况下，将II、VI、VIIa、B、VIII和XII阶段的小管的2毫米片段培养1小时。数据（每小时合成35 S-CP-2）表明CP-2的生物合成速率从阶段II到阶段VI和VIIa，B增加9倍，然后到阶段XII降低13倍。为了确定这些生物合成速率是否与分泌速率相同，将小管与[35 S]甲硫氨酸一起培养17小时，从培养基中免疫沉淀CP-2，并将数据表示为每小时分泌的35 S-CP-2。该分析表明，CP-2的分泌速率以与其合成速率相同的阶段特异性方式变化。然而，在每个阶段，该分子的表观生物合成速率超过其表观分泌速率。为了解释这一观察结果，我们分析了新合成的CP-2从小管中输出的速率。这表明，定量输出到培养基中的蛋白质需要至少17小时。这段时间最有可能是由于蛋白质的保留在管腔内，因为原代培养的支持细胞显示快速分泌新合成的CP-2。因此，我们得出结论，CP-2是以阶段依赖性方式生物合成的，并且所有CP-2都是分泌的。
Cyclic protein-2 (CP-2) is secretedin vitroin substantial amounts by mature rat Sertoli cells in intact Stage VI and Stage VII seminiferous tubules. This stage-dependent secretion has led us to postulate that the biosynthesis of this molecule is stimulated by germ cells at a specific state of development. In order to explore this hypothesis and to examine the steps in CP-2's biosynthesis, we generated a polyclonal antiserum against this protein and used it to analyze the biosynthesis and secretion of CP-2. Analysis of the steps in the biosynthesis of CP-2 indicated that its polypeptide core represented most if not all of the translation product of the CP-2 mRNA and that a single asparginelinked oligosaccharide became attached to this core. Analysis of the rate of biosynthesis of CP-2 at specific stages of the cycle of the seminiferous epithelium was also conducted. Two-millimeter segments of tubules at Stages II, VI, VIIa, b, VIII, and XII were cultured for 1 hr in the presence of [35S]methionine and radiolabeled CP-2 immunoprecipitated from the tubules. Data (35S-CP-2 synthesized per hour) demonstrated that the rate of CP-2's biosynthesis increased 9-fold from Stage II to Stages VI and VIIa, b and then decreased 13-fold by Stage XII. To determine whether these rates of biosynthesis were identical to the rates of secretion, tubules were cultured for 17 hr with [35S]methionine, CP-2 was immunoprecipitated from the culture medium and data were expressed as35S-CP-2 secreted per hour. This analysis demonstrated that the rate of secretion of CP-2 varied in the same stage-specific manner as its rate of synthesis. However, at each stage, the apparent rate of biosynthesis of the molecule exceeded its apparent rate of secretion. In order to explain this observation, we analyzed the rate of export of newly synthesized CP-2 out of the tubules. This demonstrated that quantitative export of the protein into culture medium required at least 17 hr. This period of time was most likely due to the retention of the protein within the tubular lumen, since primary cultures of Sertoli cells were shown to rapidly secrete newly synthesized CP-2. We, therefore, concluded that CP-2 was biosynthesized in a stage-dependent manner and that all CP-2 was secreted.