Oppenheimer C L, Pessin J E, Massague J, Gitomer W, Czech M P
J Biol Chem. 1983 Apr 25;258(8):4824-30.
Incubation of intact rat adipocytes with physiological concentrations of insulin stimulates binding of insulin-like growth factor II (IGF-II) to its receptor by 3- to 10-fold. The effect is temperature- and dose-dependent, with 0.1 nM insulin giving half-maximal stimulation. Scatchard analysis of IGF-II binding to intact adipocytes indicates that this effect is due to an apparent increase in receptor affinity, from Kd = 63 nM in the absence of insulin to Kd = 5.8 nM in the presence of 10 nM insulin, with no apparent change in the number of cell surface binding sites (220,000/cell). Scatchard analysis of 125I-IGF-II binding to isolated membrane fractions demonstrated that all IGF-II receptors in plasma membranes and low density microsomes from control cells are converted during homogenization to the high affinity form (Kd = 2 to 6 nM) seen in insulin-treated intact adipocytes. No significant difference in affinity was observed between plasma membranes from control or insulin-treated adipocytes or between low density microsomes from control or insulin-treated cells. However, in apparent contrast to the results obtained in intact adipocytes, the number of binding sites is increased in the plasma membrane fraction from insulin-treated cells by an average of 60%, while the number of receptors is decreased by 40% in low density microsomes from insulin-treated cells compared to control cells. These results were confirmed by direct visualization of the Mr = 270,000 IGF-II receptor band on dodecyl sulfate gels following affinity labeling with 125I-IGF-II and the cross-linker disuccinimidyl suberate. Scatchard analysis of the total cellular membranes showed no difference in the total number of binding sites between control and insulin-treated cells. These results demonstrate that insulin has two effects on the IGF-II receptor in adipocytes. 1) It rapidly increases the apparent affinity of the receptor in the intact cell without changing the apparent number of receptors on the cell surface; and 2) it induces a redistribution of the high affinity IGF-II receptor between plasma membranes and low density microsomes upon homogenization of cells and preparation of membranes. The latter effect closely parallels the insulin-induced membrane redistribution of the glucose transporter that occurs in the rat adipocyte by an unknown mechanism.
用生理浓度的胰岛素孵育完整的大鼠脂肪细胞，可使胰岛素样生长因子II（IGF-II）与其受体的结合增加3至10倍。该效应具有温度和剂量依赖性，0.1 nM胰岛素可产生半数最大刺激作用。对完整脂肪细胞中IGF-II结合的Scatchard分析表明，这种效应是由于受体亲和力明显增加，从无胰岛素时的Kd = 63 nM增加到存在10 nM胰岛素时的Kd = 5.8 nM，而细胞表面结合位点的数量没有明显变化（220,000个/细胞）。对125I-IGF-II与分离的膜组分结合的Scatchard分析表明，对照细胞的质膜和低密度微粒体中的所有IGF-II受体在匀浆过程中都转化为在胰岛素处理的完整脂肪细胞中所见的高亲和力形式（Kd = 2至6 nM）。在对照或胰岛素处理的脂肪细胞质膜之间或对照或胰岛素处理的细胞的低密度微粒体之间未观察到亲和力的显著差异。然而，与在完整脂肪细胞中获得的结果明显不同的是，胰岛素处理细胞的质膜组分中结合位点的数量平均增加了60%，而与对照细胞相比，胰岛素处理细胞的低密度微粒体中受体的数量减少了40%。在用125I-IGF-II和交联剂辛二酸二琥珀酰亚胺酯进行亲和标记后，通过十二烷基硫酸盐凝胶上Mr = 270,000的IGF-II受体条带的直接可视化证实了这些结果。对总细胞膜的Scatchard分析表明，对照细胞和胰岛素处理细胞之间结合位点的总数没有差异。这些结果表明胰岛素对脂肪细胞中的IGF-II受体有两种作用。1）它在完整细胞中迅速增加受体的表观亲和力，而不改变细胞表面受体的表观数量；2）在细胞匀浆和膜制备过程中，它诱导高亲和力IGF-II受体在质膜和低密度微粒体之间重新分布。后一种效应与胰岛素诱导的大鼠脂肪细胞中葡萄糖转运体的膜重新分布密切相似，其发生机制尚不清楚。
