Niimi S, Yamaguchi T, Hayakawa T
Division of Biological Chemistry and Biologicals, National Institute of Health Sciences, Tokyo, Japan.
J Steroid Biochem Mol Biol. 1997 Apr;61(1-2):65-71. doi: 10.1016/s0960-0760(97)00005-8.
The participation of tyrosine kinase in the regulation of the glucocorticoid receptor (GR) was studied in primary cultured rat hepatocytes using the tyrosine kinase inhibitor herbimycin A. Herbimycin A decreased the number of high-affinity binding sites of glucocorticoids in the cytosolic fraction and increased the equilibrium dissociation constant (Kd). Western blot analysis revealed that it also decreased the amount of GR protein. On the other hand, cycloheximide did not affect the GR protein level. Although herbimycin A slightly increased the amount of GR protein in the nuclear fraction, the increase was much lower than that of its decrease in the cytosolic fraction. Therefore, the decrease of GR protein in the cytosolic fraction was not caused by the inhibition of GR protein synthesis nor the translocation of GR from cytosol to nuclei. As herbimycin A also suppressed the dexamethasone (Dex)-dependent induction of tyrosine aminotransferase (TAT) activity, the decrease of GR protein was followed by the suppression of the GR-mediated biological response. These findings indicate that tyrosine kinase is necessary for the maintenance of the level of GR protein and its affinity of binding sites in the cytosolic fraction. Our results also suggested that the increase of GR protein stability is the most probable explanation for the maintenance of its level.
利用酪氨酸激酶抑制剂赫曲霉素A，在原代培养的大鼠肝细胞中研究了酪氨酸激酶在糖皮质激素受体（GR）调节中的作用。赫曲霉素A降低了胞质部分糖皮质激素高亲和力结合位点的数量，并增加了平衡解离常数（Kd）。蛋白质免疫印迹分析表明，它还降低了GR蛋白的量。另一方面，放线菌酮不影响GR蛋白水平。虽然赫曲霉素A使核部分的GR蛋白量略有增加，但增加幅度远低于其在胞质部分的减少幅度。因此，胞质部分GR蛋白的减少不是由GR蛋白合成的抑制或GR从胞质向细胞核的转运引起的。由于赫曲霉素A也抑制了地塞米松（Dex）依赖性酪氨酸转氨酶（TAT）活性的诱导，GR蛋白的减少伴随着GR介导的生物学反应的抑制。这些发现表明，酪氨酸激酶对于维持胞质部分GR蛋白水平及其结合位点的亲和力是必需的。我们的结果还表明，GR蛋白稳定性的增加是维持其水平的最可能解释。
