CULTURED ANIMAL-CELLS EXPOSED TO AMINO-ACID-ANALOGS OR PUROMYCIN RAPIDLY SYNTHESIZE SEVERAL POLYPEPTIDES
CULTURED ANIMAL-CELLS EXPOSED TO AMINO-ACID-ANALOGS OR PUROMYCIN RAPIDLY SYNTHESIZE SEVERAL POLYPEPTIDES
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DOI：
10.1002/jcp.1041020315
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发表时间：
1980-01-01
影响因子：
5.6
通讯作者：
HIGHTOWER, LE
中科院分区：
文献类型：
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作者：
HIGHTOWER, LE
四种主要的酸性多肽[P]，分子量分别为88，000、72，000、71，000和23，000，以及分子量分别为110，000、50，000、38，000和30，000的次要多肽在培养的鸡胚（CE）细胞中迅速积累，这些细胞暴露于Arg类似物刀豆氨酸3小时。P110、P88、P71、72和P23具有独特的肽图。获得了P23的27，000道尔顿前体的证据。类似物刺激的蛋白质与另一组被称为葡萄糖调节蛋白的诱导性禽类多肽无关。在哺乳动物细胞[人成纤维细胞FS-4细胞和小鼠成纤维细胞3 T3细胞]中，刀豆氨酸处理后，几种大小与鸟类蛋白相似的多肽的积累速率急剧增加。与类似物刺激的蛋白质相同的电泳迁移率，等电点和肽图的蛋白质在未经处理的文化中以低水平表达。为了确定类似物处理后刀豆氨酸介导的蛋白质积累增加和蛋白质代谢恢复的时间过程，从CE细胞中提取放射性标记的蛋白质，并在十二烷基硫酸钠聚丙烯酰胺凝胶上进行分析。在暴露于刀豆氨酸的培养物中，P88和P71，72的积累速率在apx中从基础水平增加到新的平台水平。1.5而P23需要约2.5h。当加入类似物时，放线菌素D和虫草素阻断了蛋白质积累的增加。这些抑制剂还阻断了去除刀豆氨酸后发生的增强蛋白质积累速率的快速下降。对增强蛋白质的代谢稳定性的研究表明，其积累的变化是由其合成速率的改变引起的。因此，类似物介导的反应满足诱导真核基因表达的几个标准。氨基酸类似物p-氟苯丙氨酸和氨基酰基-tRNA嘌呤霉素的链终止类似物刺激刀豆氨酸诱导的同一组蛋白质的合成。这些蛋白质的合成增强似乎是对异常蛋白质和嘌呤霉素肽的催化剂存在的细胞反应。
Four major acidic polypeptides [P], with MW of 88,000, 72,000, 71,000 and 23,000 and minor polypeptides with MW of 110,000, 50,000, 38,000 and 30,000 rapidly accumulated in cultured chick embryo (CE) cells which were exposed for 3 h to the Arg analog canavanine. P110, P88, P71,72, and P23 had unique peptide maps. Evidence of a 27,000 dalton precursor to P23 was obtained. The analog-stimulated proteins were not related to another set of inducible avian polypeptides known as the glucose-regulated proteins. In mammalian cells [human fibroblast FS-4 cells and mouse fibroblast 3T3 cells], the rate of accumulation of several polypeptides, which were similar in size to the avian proteins, sharply increased after canavanine treatment. Proteins with the same electrophoretic mobilities, isoelectric points and peptide maps as the analog-stimulated proteins were expressed at low levels in untreated cultures. To determine the time courses of the canavanine-mediated increases in protein accumulation and the recovery of protein metabolism after analog treatment, radioactively labeled proteins were extracted from CE cells and analyzed on sodium dodecyl sulfate polyacrylamide gels. In cultures exposed to canavanine, the rates of accumulation of P88 and P71,72 increased from basal to new plateau levels in .apprx. 1.5 h, while P23 required about 2.5 h. When added with the analog, actinomycin D and cordycepin blocked the increases in protein accumulation. These inhibitors also blocked the rapid decline in the rates of accumulation of the enhanced proteins which occurred after removal of canavanine. Studies of the metabolic stability of the enhanced proteins indicated that the changes in their accumulation were caused by alterations in their rates of synthesis. Thus, the analog-mediated response fulfilled several of the criteria for inducible eucaryotic gene expression. The amino acid analog p-fluorophenylalanine and the chain-terminating analog of amino acyl-tRNA puromycin stimulated the synthesis of the same set of proteins induced by canavanine. The enhanced synthesis of these proteins appeared to be a cellular response to either the presence of catabolism of abnormal proteins and puromycyl peptides.