Effect of growth hormone on protein phosphorylation in isolated rat hepatocytes.
Effect of growth hormone on protein phosphorylation in isolated rat hepatocytes.
复制标题
生长激素对离体大鼠肝细胞蛋白质磷酸化的影响。
DOI：
10.1021/bi00377a009
复制
发表时间：
1987
期刊：
影响因子：
2.9
通讯作者：
Donner,DB
中科院分区：
文献类型：
--
作者：
Yamada,K;Lipson,KE;Marino,MW;Donner,DB
修订稿于 1986 年 10 月 20 日收到 摘要：雄性大鼠的肝细胞与 [32P] P 一起孵育； 37°C 40 分钟，从而用 32P 平衡细胞 ATP 池。随后再暴露于牛生长激素 10 分钟并没有改变细胞 [t-32P] ATP 的比活性。使用二维凝胶电泳或色谱聚焦，然后进行十二烷基硫酸钠-聚丙烯酰胺凝胶电泳来分级从对照或激素刺激的细胞中溶解的磷蛋白。在37℃下用5nM生长激素刺激肝细胞10分钟影响许多蛋白质的磷酸化，包括p/4.7的Mr 46000种，其磷酸化增强了(2.65±0.50)倍。生长激素对 MT 46 000 物种磷酸化的最大影响的很大一部分是由 1-5% 的受体占据引起的。与体细胞受体高度结合的牛生长激素，或未受其他激素污染的重组人生长激素，也同样影响肝蛋白的磷酸化。细胞匀浆离心后，Mr 46 000 磷蛋白被分离到富含细胞质的级分中。胰岛素和胰高血糖素也分别使 Mx 46 000 磷酸蛋白的磷酸化增加 (1.75±0.35) 倍和 (2.15±0.50) 倍。这些观察结果与细胞蛋白磷酸化状态的选择性变化可能介导细胞中生长激素作用的可能性是一致的。导致生长激素作用的第一步是与受体结合，这些受体已在许多不同的细胞和细胞膜中得到鉴定（Kelly 等人，1974 年；Lesniak 等人，
Revised Manuscript Received October 20, 1986 abstract: Hepatocytes from male rats were incubated with [32P] P; for 40 min at 37 C, thereby equilibrating the cellular ATP pool with 32P. Subsequent exposure to bovine growth hormone for 10 additional min did not change the specific activity of cellular [t-32P] ATP. Two-dimensional gel electrophoresis or chromatofocusing followed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis was used to fractionate phosphoproteins solubilizedfrom control or hormone-stimulated cells. Stimulation of hepatocytes with 5 nM growth hormone for 10 min at 37 C affected the phosphorylation of a number of proteins including an Mr 46000 species of p/4.7 whose phosphorylation was augmented (2.65±0.50)-fold. A significant fraction of the maximal effect of growth hormone on phosphorylation of the MT 46 000 species was elicited by 1-5% receptor occupancy. Bovine growth hormone, which binds to somatogenic receptors with great specificity, or recombinant human growth hormone, which is not contaminated with other hormones, affected phosphorylation of hepatic proteins similarly. The Mr 46 000phosphoprotein was isolated in a fraction enriched in cytosol after centrifugation of cellular homogenates. Phosphorylation of the Mx 46 000 phos-phoprotein was also increased (1.75±0.35)-fold and (2.15±0.50)-fold by insulin and glucagon, respectively. These observations are consistent with the possibilitythat selective changes in the phosphorylation state of cellular proteins may mediate growth hormone actions in cells. e first step leading to growth hormone action is binding to receptors, which have been identified in a number of dif-ferent cells and membranes (Kelly et al., 1974; Lesniak et al.,