Katze M G, DeCorato D, Krug R M
J Virol. 1986 Dec;60(3):1027-39. doi: 10.1128/JVI.60.3.1027-1039.1986.
During influenza virus infection, protein synthesis is maintained at high levels and a dramatic switch from cellular to viral protein synthesis occurs despite the presence of high levels of functional cellular mRNAs in the cytoplasm of infected cells (M. G. Katze and R. M. Krug, Mol. Cell. Biol. 4:2198-2206, 1984). To determine the step at which the block in cellular mRNA translation occurs, we compared the polysome association of several representative cellular mRNAs (actin, glyceraldehyde-3-phosphate dehydrogenase, and pHe7 mRNAs) in infected and uninfected HeLa cells. We showed that most of these cellular mRNAs remained polysome associated after influenza viral infection, indicating that the elongation of the proteins encoded by these cellular mRNAs was severely inhibited. Because the polysomes containing these cellular mRNAs did not increase in size but either remained the same size or decreased in size, the initiation step in cellular protein synthesis must also have been defective. Several control experiments established that the cellular mRNAs sedimenting in the polysome region of sucrose gradients were in fact associated with polyribosomes. Most definitively, puromycin treatment of infected cells caused the dissociation of polysomes and the release of cellular, as well as viral, mRNAs from the polysomes, indicating that the cellular mRNAs were associated with polysomes that were capable of forming at least a single peptide bond. A similar analysis was performed with HeLa cells infected by adenovirus, which also dramatically shuts down cellular protein synthesis. Again, it was found that most of the cellular mRNAs, which were translatable in reticulocyte extracts, remained associated with polysomes and that there was a combined initiation-elongation block to cellular protein synthesis. In cells infected by both adenovirus and influenza virus, influenza viral mRNAs were on larger polysomes than were several late adenoviral mRNAs with comparably sized coding regions. In addition, after influenza virus superinfection of cells infected by the adenovirus mutant dl331, a situation in which there is a limitation in the amount of functional initiation factor eIF-2 (M. G. Katze, B. M. Detjen, B. Safer, and R. M. Krug, Mol. Cell. Biol. 6:1741-1750, 1986), influenza viral mRNAs, but not late adenoviral mRNAs, were on polysomes. These results indicate that influenza viral mRNAs are better initiators of translation than are late adenoviral mRNAs.
在流感病毒感染期间，尽管被感染细胞的细胞质中存在高水平的功能性细胞mRNA，但蛋白质合成仍维持在高水平，并且发生了从细胞蛋白质合成到病毒蛋白质合成的显著转变（M.G.卡茨和R.M.克鲁格，《分子细胞生物学》4:2198 - 2206，1984年）。为了确定细胞mRNA翻译受阻发生的步骤，我们比较了感染和未感染的HeLa细胞中几种代表性细胞mRNA（肌动蛋白、甘油醛 - 3 - 磷酸脱氢酶和pHe7 mRNA）与多核糖体的结合情况。我们发现，在流感病毒感染后，这些细胞mRNA中的大多数仍与多核糖体结合，这表明这些细胞mRNA编码的蛋白质的延伸受到了严重抑制。由于含有这些细胞mRNA的多核糖体大小没有增加，而是保持不变或减小，细胞蛋白质合成的起始步骤也必定存在缺陷。几个对照实验证实，在蔗糖梯度的多核糖体区域沉降的细胞mRNA实际上与多聚核糖体相关。最明确的是，用嘌呤霉素处理感染细胞会导致多核糖体解离，并使细胞和病毒mRNA从多核糖体上释放出来，这表明细胞mRNA与能够形成至少一个肽键的多核糖体相关。对感染腺病毒的HeLa细胞进行了类似分析，腺病毒感染也会显著关闭细胞蛋白质合成。同样发现，大多数在网织红细胞提取物中可翻译的细胞mRNA仍与多核糖体结合，并且细胞蛋白质合成存在起始 - 延伸联合阻断现象。在同时感染腺病毒和流感病毒的细胞中，流感病毒mRNA所在的多核糖体比具有相当大小编码区域的几种晚期腺病毒mRNA所在的多核糖体更大。此外，在用腺病毒突变体dl331感染的细胞中进行流感病毒超感染后（在这种情况下功能性起始因子eIF - 2的量有限，M.G.卡茨、B.M.德特延、B.萨弗和R.M.克鲁格，《分子细胞生物学》6:1741 - 1750，1986年），流感病毒mRNA存在于多核糖体上，而晚期腺病毒mRNA则不存在。这些结果表明，流感病毒mRNA比晚期腺病毒mRNA是更好的翻译起始者。
