van Deurs B, Petersen O W, Olsnes S, Sandvig K
Exp Cell Res. 1987 Jul;171(1):137-52. doi: 10.1016/0014-4827(87)90257-6.
Galactose-terminating membrane glycoproteins and glycolipids on two established human breast carcinoma cell lines were tagged at 4 degrees C with a ricin-horseradish peroxidase conjugate (Ri-HRP). The cells were then incubated for various periods of time at 37 or 18 degrees C. After fixation and diaminobenzidine cytochemistry, the compartments reached by Ri-HRP were studied by analyzing thin serial sections. In both cell types a highly pleomorphic endosomal system comprising vacuolar elements as well as smaller, sometimes branched, tubular elements (tubular endosomes) was revealed at both 37 and 18 degrees C. At 37 degrees C Ri-HRP was consistently observed in flattened cisterns of the Golgi region in 30-40% of the Golgi complexes examined after 30-60 min of incubation. However, no Ri-HRP reached such Golgi elements at 18 degrees C, even after incubation for 180 min. Moreover, at 18 degrees C the ability of ricin to inhibit protein synthesis was virtually abolished, whereas the effect of diphtheria toxin was reduced much less. Following incubation with a monovalent transferrin-HRP conjugate or with unconjugated HRP, no labeling of cisternal Golgi elements was detected. These data indicate that delivery of galactose-terminating membrane molecules from endosomes to the Golgi complex is a discontinuous, temperature-sensitive process and that this process may be required for optimal ricin A-chain translocation.
在4℃下，用蓖麻毒素 - 辣根过氧化物酶偶联物（Ri - HRP）标记两种已建立的人乳腺癌细胞系上的半乳糖末端膜糖蛋白和糖脂。然后将细胞在37℃或18℃下孵育不同时间。固定并进行二氨基联苯胺细胞化学后，通过分析连续薄切片研究Ri - HRP到达的区室。在37℃和18℃下，两种细胞类型均显示出高度多形性的内体系统，包括液泡成分以及较小的、有时分支的管状成分（管状内体）。在37℃下，孵育30 - 60分钟后，在30 - 40%的高尔基体复合物的扁平潴泡中持续观察到Ri - HRP。然而，在18℃下，即使孵育180分钟，也没有Ri - HRP到达这样的高尔基体成分。此外，在18℃下，蓖麻毒素抑制蛋白质合成的能力几乎被消除，而白喉毒素的作用降低程度要小得多。用单价转铁蛋白 - HRP偶联物或未偶联的HRP孵育后，未检测到高尔基体潴泡成分的标记。这些数据表明，从内体向高尔基体复合物递送半乳糖末端膜分子是一个不连续的、温度敏感的过程，并且这个过程可能是蓖麻毒素A链最佳易位所必需的。
