Smith C J, Wejksnora P J, Warner J R, Rubin C S, Rosen O M
Proc Natl Acad Sci U S A. 1979 Jun;76(6):2725-9. doi: 10.1073/pnas.76.6.2725.
A protein of molecular weight 31,000 became labeled with 32P within 5 min after addition of insulin to differentiated 3T3-L1 preadipocytes previously incubated for 55 min with 32Pi. The effect was mimicked by antisera directed against the insulin receptor and was eliminated by anti-insulin antiserum. Incorporation of 32P into this protein was more than 20-fold greater in insulin-treated cells than in cells not exposed to the hormone. At concentrations greater than required with insulin, epidermal growth factor and serum (1-5%) also stimulated phosphorylation whereas 1-isoproterenol, a beta-adrenergic agonist that increases intracellular accumulation of cyclic AMP, was without effect. The 31,000-dalton protein has been tentatively identified as ribosomal protein S6 by two-dimensional polyacrylamide gel electrophoresis. Incorporation of 32P into S6 could be detected within the same time period (5 min) and at the same insulin concentrations (0.1-1.0 nM) as are required to stimulate hexose uptake in both 3T3-L1 cells and mature mammalian adipocytes. The mechanism by which this phosphorylation either mediates or reflects the intracellular actions of insulin remains to be elucidated.
在向预先用32Pi孵育55分钟的分化3T3-L1前脂肪细胞中添加胰岛素后5分钟内，一种分子量为31,000的蛋白质被32P标记。针对胰岛素受体的抗血清模拟了这种效应，而抗胰岛素抗血清消除了这种效应。在胰岛素处理的细胞中，该蛋白质中32P的掺入量比未暴露于该激素的细胞中高20倍以上。在高于胰岛素所需浓度时，表皮生长因子和血清（1-5%）也刺激磷酸化，而增加细胞内环磷酸腺苷细胞内积累的β-肾上腺素能激动剂1-异丙肾上腺素则无作用。通过二维聚丙烯酰胺凝胶电泳，暂定将这种31,000道尔顿的蛋白质鉴定为核糖体蛋白S6。在刺激3T3-L1细胞和成熟哺乳动物脂肪细胞摄取己糖所需的相同时间段（5分钟）和相同胰岛素浓度（0.1-1.0 nM）下，可以检测到32P掺入S6。这种磷酸化介导或反映胰岛素细胞内作用的机制仍有待阐明。
