Microtubules are acetylated in domains that turn over slowly.
Microtubules are acetylated in domains that turn over slowly.
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发表时间：
1989
影响因子：
4
通讯作者：
D. R. Webster;G. Borisy
中科院分区：
文献类型：
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作者：
D. R. Webster;G. Borisy
微管蛋白受到翻译后乙酰化反应的影响，这被认为与修饰微管（mt）的稳定性增加有关。我们试图通过确定其特定的周转率来直接测试乙酰化mt的稳定性。我们使用了人类成纤维细胞，它包含一个MTs子集，显示乙酰化的末端和内部结构域。通过向细胞微注射生物素化的脑微管蛋白来分析乙酰化结构域的转换，并通过三标记免疫荧光测定生物素化的脑微管蛋白进入乙酰化和非乙酰化结构域的进展。注射后2分钟内，生物素化结构域几乎与所有观察到的非乙酰化MT末端相邻，但与末端乙酰化结构域不相邻，表明前者在生长，后者没有生长。注射10分钟后，许多缺乏乙酰化结构域的mt均匀地结合了生物素化亚基，而大多数含有乙酰化结构域的mt仍然未标记，这表明非乙酰化的mt正在翻转，而大多数乙酰化结构域则没有。注射一小时后，几乎所有非乙酰化的mt都被标记为生物素，而大约一半的乙酰化结构域含有生物素，这表明乙酰化结构域的翻转速度比非乙酰化的大阵列慢得多。乙酰化结构域两侧的非乙酰化MT区域也缺乏半抗原，这表明乙酰化修饰了稳定MT的离散区域。注射后16小时，未进入有丝分裂的细胞仍保留未翻动的乙酰化结构域（占所有乙酰化结构域的13%），表明乙酰化结构域可以非常长寿。（摘要删节250字）
Tubulin is subject to a post-translational acetylation reaction that is thought to be correlated with increased stability of the modified microtubules (MTs). We sought to test directly the stability of acetylated MTs by determining their specific rate of turnover. We used human fibroblasts, which contain a subset of MTs that display terminal and internal domains of acetylation. The turnover of acetylated domains was analysed by microinjecting cells with biotinylated brain tubulin and determining, by triple-label immunofluorescence, the progress of incorporation of biotinylated tubulin into acetylated and non-acetylated domains. Within two minutes after injection, biotinylated domains were contiguous with virtually all observed non-acetylated MT ends but were not contiguous with terminal acetylated domains, demonstrating that the former were growing while the latter were not. Ten minutes after injection, many MTs lacking acetylated domains had incorporated biotinylated subunits uniformly while most MTs containing acetylated domains remained unlabelled, indicating that non-acetylated MTs were turning over while most acetylated domains were not. One hour after injection, virtually all non-acetylated MTs were labelled with biotin whereas approximately half of the acetylated domains contained biotin, demonstrating that acetylated domains turned over much more slowly than the non-acetylated, bulk array. Non-acetylated MT regions flanking acetylated domains also lacked hapten, indicating that acetylation modified discrete regions along stable MTs. Sixteen hours after injection, cells that had not entered mitosis still retained acetylated domains that had not turned over (13% of all acetylated domains), indicating that acetylated domains can be extremely long-lived.(ABSTRACT TRUNCATED AT 250 WORDS)