TUBULIN MESSENGER-RNA INSTABILITY AND STABILIZATION BY PROTEIN-SYNTHESIS INHIBITORS ARE REPRODUCIBLE IN NONTRANSLATING EXTRACTS FROM CHLAMYDOMONAS
TUBULIN MESSENGER-RNA INSTABILITY AND STABILIZATION BY PROTEIN-SYNTHESIS INHIBITORS ARE REPRODUCIBLE IN NONTRANSLATING EXTRACTS FROM CHLAMYDOMONAS
复制标题
DOI：
10.1002/dvg.1020140607
复制
发表时间：
1993-01-01
期刊：
影响因子：
--
通讯作者：
BAKER, EJ
中科院分区：
文献类型：
--
作者：
BAKER, EJ
在莱茵衣藻中，鞭毛截除会刺激鞭毛蛋白的合成，从而使细胞能够快速再生鞭毛。诱导涉及大量 mRNA（包括编码微管蛋白的 mRNA）的协调积累和快速降解。诱导后的微管蛋白 mRNA 的诱导后降解已被证明在两个方面与组成型周转途径不同：(1) 降解速率加快，(2) 通过抑制蛋白质合成来防止降解。在本报告中，表明放线菌酮 (CX) 可以阻止所有爆破诱导的 mRNA 的诱导后降解，这表明它们都可能通过相同的途径降解。已开发出无细胞腐烂系统来研究降解途径。在这些提取物中，微管蛋白 mRNA 降解的至少两个特征是可重现的：(1) 内源性 α-微管蛋白 mRNA 的稳定性低于同一提取物中的组成型 mRNA，以及 (2) 从 CX 处理的细胞制备的提取物（CX 提取物）中的 α-微管蛋白 mRNA 明显比未处理细胞的提取物（对照提取物）中的更稳定。这表明 CX 阻止体内微管蛋白 mRNA 快速降解的机制不仅仅是阻止其翻译，而且表明涉及改变的反式因子。当提取物在核糖体与 mRNP 解离的条件下制备时，两种提取物中微管蛋白 mRNA 稳定性的差异得以维持，表明完整的多核糖体结构不是必需的。从对照和 CX 提取物中分离出的含有微管蛋白 mRNA 的多核糖体在单独测定时同样稳定。然而，来自对照提取物的多核糖体比来自 CX 提取物的多核糖体对外源 RNAse 处理更敏感，表明结构差异。对照和 CX 提取物之间影响微管蛋白 mRNA 降解率的可溶性因子没有可检测到的差异；向对照或 CX 提取物中添加过量的可溶性因子不会改变提取物中微管蛋白 mRNA 的降解，两种提取物的简单一对一组合也不会导致混合物中整个微管蛋白 mRNA 群体的稳定或不稳定。作为一个整体，去鞭毛诱导的 mRNA 对提取物中的核酸酶活性特别敏感，可被氧钒核糖核苷复合物抑制，而该复合物似乎不会攻击组成型 mRNA。有人提出，在存在和不存在 CX 的情况下产生的微管蛋白 mRNP 的结构差异是其稳定性差异的基础，并且常见的核酸酶靶向诱导的鞭毛蛋白 mRNA。 (C) 1993 Wiley-Liss, Inc.
In Chlamydomonas reinhardtii, flagellar amputation stimulates an induction in the synthesis of flagellar proteins which allows the cells to rapidly regenerate their flagella. The induction involves the coordinate accumulation and rapid degradation of a large number mRNAs, including those encoding the tubulins. The post-induction degradation of induced tubulin mRNAs has been shown to differ from the constitutive turnover pathway in two ways: (1) the rate of degradation is accelerated, and (2) degradation is prevented by inhibition of protein synthesis. In this report, it is shown that the post-induction degradation of all deflagellation-induced mRNAs era mined is prevented by cycloheximide (CX), suggesting they all may be degraded via the same pathway. A cell-free decay system has been developed to investigate the degradation pathway. At least two characteristics of tubulin mRNA degradation are reproducible in these extracts: (1) endogenous alpha-tubulin mRNA is less stable than constitutive mRNAs in the same extract and (2) alpha-tubulin mRNA in extracts prepared from CX-treated cells (CX extracts) is significantly more stable than it is in extracts from untreated cells (control extracts). This indicates that the mechanism by which CX blocks rapid degradation of tubulin mRNA in vivo is not simply by preventing its translation and suggests the involvement of an altered trans-factor. The difference in tubulin mRNA stability in the two extracts is maintained when the extracts are prepared under conditions that dissociate ribosomes from mRNPs, indicating intact polysome structure is not necessary. Tubulin mRNA-containing polysomes isolated from control and CX extracts are equally stable when assayed alone. However, the polysomes from control extracts are more sensitive to exogenous RNAse treatment than are those from CX extracts, indicating a structural difference. There are no detectable differences in soluble factors that influence tubulin mRNA degradation rate between control and CX extracts; addition of excess soluble factors to either control or CX extracts does not alter the tubulin mRNA degradation in the extract, nor does a simple one-to-one combination of the two extracts result in stabilization or destabilization of the whole population of tubulin mRNAs in the mixture. The deflagellation-induced mRNAs, as a group, are shown to be particularly susceptible to a nuclease activity in extracts, inhibitable by vanadyl ribonucleoside complexes, which does not appear to attack constitutive mRNAs. It is proposed that a structural difference in the tubulin mRNPs produced in the presence and absence of CX underlies their differences in stabilities, and that a common nuclease targets the induced flagellar protein mRNAs. (C) 1993 Wiley-Liss, Inc.