Translational control of phage f1 gene expression by differential activities of the gene V, VII, IX and VIII initiation sites.
Translational control of phage f1 gene expression by differential activities of the gene V, VII, IX and VIII initiation sites.
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通过基因 V、VII、IX 和 VIII 起始位点的差异活性对噬菌体 f1 基因表达进行翻译控制。
DOI：
10.1016/0022-2836(87)90557-2
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发表时间：
1987
影响因子：
5.6
通讯作者：
Steege,DA
中科院分区：
文献类型：
--
作者：
Blumer,KJ;Ivey,MR;Steege,DA
在大肠杆菌感染丝状噬菌体（f1, M13, fd）后，噬菌体特异性转录和随后的RNA加工产生了丰富且寿命相对较长的噬菌体mRNA，编码4个相邻基因V， VII， IX和VIII。然而，基因V和基因VIII的产物的合成水平远高于基因VII和基因IX的蛋白质。为了了解这些基因的翻译起始位点是否在活性和/或功能特性上表现出相应的差异，我们从感染f1的细胞中纯化了一些噬菌体mrna，并在体外起始反应中对它们进行了检测。获得的噬菌体mRNA种类的核糖体结合模式和包含选定起始区域的较小定义RNA片段的核糖体结合模式显示了大范围的表观核糖体结合强度。基因V和基因VIII位点在它们存在的每个mRNA物种中都被有效识别。基因IX位点的活性似乎受到局部mRNA结构的限制：该位点在所有噬菌体mRNA物种中都无法检测到或核糖体结合活性低，但如果从启动物AUG上游15个核苷酸处形成潜在发夹茎环所需的RNA序列被移除，其活性至少提高10倍。基因VII位点未显示出与任何噬菌体mRNA或RNA片段核糖体相互作用的证据。通过克隆含有基因V或基因VII起始位点序列的f1 DNA片段来驱动β-半乳糖苷酶合成，在体内观察到同样显著的起始活性差异。高水平的基因V-β-半乳糖苷酶融合蛋白在V位点启动，但在VII位点没有可检测到的合成。然而，如果在VII位点之前有上游基因V的所有编码信息，则会产生少量在VII位点启动的融合蛋白。总体结果与噬菌体感染细胞中观察到的蛋白质产量一致，提供了强有力的证据，证明这些翻译起始位点的特性在很大程度上决定了噬菌体f1基因V、VII、IX和VIII的差异表达。
Phage-specific transcription and subsequent RNA processing inEscherichia coliinfected with the filamentous phage (f1, M13, fd) generate a pool of abundant and relatively long-lived phage mRNA species encoding the four adjacent genes V, VII, IX and VIII. Yet the products of gene V and gene VIII are synthesized at much higher levels than the gene VII and gene IX proteins. To ask if the translational initiation sites heading these genes show corresponding differences in activity and/or functional properties, we have purified a number of the phage mRNAs from cells infected with f1 and examined them inin vitroinitiation reactions. The ribosome binding patterns obtained for the phage mRNA species and for smaller defined RNA fragments containing selected initiator regions reveal a large range in apparent ribosome binding strengths. The gene V and gene VIII sites are recognized efficiently in each mRNA species in which they are present. Gene IX site activity appears to be limited by local mRNA structure: the site has undetectable or low ribosome binding activity in all of the phage mRNA species, but is at least tenfold more active if the RNA sequences required to form a potential hairpin stem-and-loop 15 nucleotides upstream from the initiator AUG have been removed. The gene VII site shows no evidence of interaction with ribosomes in any phage mRNA or RNA fragment tested. The same striking differences in initiation activity were observedin vivoby cloning small f1 DNA fragments containing gene V or gene VII initiation site sequences to drive β-galactosidase synthesis. High levels of a gene V-β-galactosidase fusion protein are initiated at the V site, but no detectable synthesis occurs from the VII site. If the VII site is preceded by all of the information encoding the upstream gene V, however, modest amounts of a fusion protein initiated at the VII site are produced. The overall results, in accord with the observed yields of proteins in the phage-infected cell, provide strong evidence that the properties of these translational initiation sites determine in a significant way the differential expression of phage f1 genes V, VII, IX and VIII.