Janszen F H, Cooke B A, van Driel M J, van der Molen H J
Biochem J. 1978 Apr 15;172(1):147-53. doi: 10.1042/bj1720147.
The amount of (35)S incorporated into the various proteins after separation by electrophoresis on sodium dodecyl sulphate/polyacrylamide gels was used as an estimate of their synthesis in the Leydig cells. Increased synthesis of proteins with apparent mol.wts. 27000 and 29000 was observed 3h after addition of lutropin to tumour Leydig cells. Incubation of Leydig cells from immature rats with lutropin (100ng/ml) for 2h or longer resulted in increased synthesis of proteins with apparent mol.wts. 11000, 21000, 27000 and 29000. At higher concentrations (>/=100ng/ml) of lutropin there was a decrease in the synthesis of a protein with apparent mol.wt. 13000. The amount of lutropin required for the stimulation of protein synthesis in both types of Leydig cells was similar to that needed for stimulation of steroidogenesis. Lutropin-stimulated specific protein synthesis was not due to increased concentrations of testosterone, however, because (1) addition of testosterone to the cells had no effect on the synthesis of the proteins, and (2) inhibition of steroidogenesis with elipten phosphate (an inhibitor of the cholesterol side-chain-cleavage enzyme complex) did not abolish the effect of lutropin. The stimulation of specific protein synthesis was also not due to contaminating follitropin in the lutropin preparation. Addition of actinomycin D to the cells at the start of the incubation prevented the effect of lutropin on specific protein synthesis, indicating that mRNA synthesis may be needed for this effect of lutropin. Incubation of the cells with cycloheximide for 30min after labelling of the proteins did not result in a detectable decrease in the amounts of the lutropin-induced proteins, indicating that their half-life is longer than 30min.
通过十二烷基硫酸钠/聚丙烯酰胺凝胶电泳分离后，掺入到各种蛋白质中的（35）S量被用作莱迪希细胞中蛋白质合成的估计值。向肿瘤莱迪希细胞中添加促黄体生成素3小时后，观察到表观分子量为27000和29000的蛋白质合成增加。用促黄体生成素（100ng/ml）孵育未成熟大鼠的莱迪希细胞2小时或更长时间，导致表观分子量为11000、21000、27000和29000的蛋白质合成增加。在较高浓度（≥100ng/ml）的促黄体生成素下，表观分子量为13000的一种蛋白质的合成减少。两种类型的莱迪希细胞中刺激蛋白质合成所需的促黄体生成素量与刺激类固醇生成所需的量相似。然而，促黄体生成素刺激的特异性蛋白质合成并非由于睾酮浓度升高，因为（1）向细胞中添加睾酮对蛋白质合成没有影响，并且（2）用磷酸依立替康（胆固醇侧链裂解酶复合物的抑制剂）抑制类固醇生成并没有消除促黄体生成素的作用。特异性蛋白质合成的刺激也不是由于促黄体生成素制剂中污染的促卵泡生成素。在孵育开始时向细胞中添加放线菌素D可阻止促黄体生成素对特异性蛋白质合成的作用，表明促黄体生成素的这种作用可能需要mRNA合成。在蛋白质标记后用环己酰亚胺孵育细胞30分钟，未导致促黄体生成素诱导的蛋白质数量出现可检测到的减少，表明它们的半衰期超过30分钟。
