Ultrastructural evidence for the accumulation of insulin in nuclei of intact 3T3-L1 adipocytes by an insulin-receptor mediated process.
Ultrastructural evidence for the accumulation of insulin in nuclei of intact 3T3-L1 adipocytes by an insulin-receptor mediated process.
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通过胰岛素受体介导的过程，胰岛素在完整 3T3-L1 脂肪细胞的细胞核中积累的超微结构证据。
DOI：
10.1073/pnas.84.2.459
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发表时间：
1987
影响因子：
11.1
通讯作者：
Jarett,L
中科院分区：
文献类型：
--
作者：
Smith,RM;Jarett,L
单体铁蛋白标记的胰岛素 (Fm-Ins) 是一种占据胰岛素受体的生物活性电子致密标记物，用于表征 3T3-L1 脂肪细胞中胰岛素的内化。 Fm-Ins 特异性结合胰岛素受体，并以时间和温度依赖性方式内化。 37°C 5-10 分钟内，Fm-Ins 在细胞质囊泡中被发现，随后在多囊泡体和溶酶体中观察到。此外，30 分钟后，少量 Fm-Ins 与细胞核结合。细胞核中观察到的 Fm-Ins 颗粒数量继续以时间依赖性方式增加，直到至少 90 分钟。在细胞核中，通常会在同一个位置（靠近核孔，与浓缩染色质的外围相关）中发现几个 Fm-Ins 颗粒。添加 250 倍过量的未标记胰岛素或在 15°C 下孵育，90 分钟后细胞核中发现的 Fm-Ins 颗粒数量分别减少了 99% 或 92%。 37℃ 90 分钟后，未标记铁蛋白的核积累仅为 Fm-Ins 的 2%。利用 125I 标记胰岛素和亚细胞分级分离的生化实验表明，完整的 3T3-L1 脂肪细胞快速内化胰岛素，1 小时后约等于 3% 的内化配体在细胞核中积累。这些数据提供了生化和高分辨率超微结构证据，表明 3T3-L1 脂肪细胞通过胰岛素受体介导的过程在细胞核中积累了潜在大量的胰岛素。胰岛素或胰岛素受体复合物向该细胞或其他细胞的细胞核的转运可能直接参与胰岛素的长期生物效应，特别是在 DNA 和 RNA 合成的控制中。
Monomeric ferritin-labeled insulin (Fm-Ins), a biologically active, electron-dense marker of occupied insulin receptors, was used to characterize the internalization of insulin in 3T3-L1 adipocytes. Fm-Ins bound specifically to insulin receptors and was internalized in a time- and temperature-dependent manner. Fm-Ins was found in cytoplasmic vesicles within 5-10 min at 37 degrees C and subsequently was observed in multivesicular bodies and lysosomes. In addition, small amounts of Fm-Ins were associated with nuclei after 30 min. The number of Fm-Ins particles observed in nuclei continued to increase in a time-dependent manner until at least 90 min. In the nucleus, several Fm-Ins particles usually were found in the same general location--near nuclear pores, associated with the periphery of the condensed chromatin. Addition of a 250-fold excess of unlabeled insulin or incubation at 15 degrees C reduced the number of Fm-Ins particles found in nuclei after 90 min by 99% or 92%, respectively. Nuclear accumulation of unlabeled ferritin was only 2% of that found with Fm-Ins after 90 min at 37 degrees C. Biochemical experiments utilizing 125I-labeled insulin and subcellular fractionation indicated that intact 3T3-L1 adipocytes internalized insulin rapidly and that approximately equal to 3% of the internalized ligand accumulated in nuclei after 1 hr. These data provide biochemical and high-resolution ultrastructural evidence that 3T3-L1 adipocytes accumulate potentially significant amounts of insulin in nuclei by an insulin receptor-mediated process. The transport of insulin or the insulin-receptor complex to nuclei in this cell or in others may be directly involved in the long-term biological effects of insulin--in particular, in the control of DNA and RNA synthesis.