TRANSLATION OF MULV AND MSV RNAS IN NUCLEASE-TREATED RETICULOCYTE EXTRACTS - ENHANCEMENT OF GAG-POL POLYPEPTIDE WITH YEAST SUPPRESSOR TRANSFER-RNA
TRANSLATION OF MULV AND MSV RNAS IN NUCLEASE-TREATED RETICULOCYTE EXTRACTS - ENHANCEMENT OF GAG-POL POLYPEPTIDE WITH YEAST SUPPRESSOR TRANSFER-RNA
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DOI：
10.1016/0092-8674(78)90149-6
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发表时间：
1978-01-01
期刊：
影响因子：
64.5
通讯作者：
GESTELAND, R
中科院分区：
文献类型：
--
作者：
PHILIPSON, L;ANDERSSON, P;GESTELAND, R
将莫洛尼小鼠白血病病毒（MuLV）和莫洛尼小鼠肉瘤病毒（MSV）的病毒粒子RNA在微球菌核酸酶处理的[兔网织细胞]无细胞系统中翻译。35S MuLV RNA形成的主要多肽分子量为78,000和65,000道尔顿，也观察到分子量为180,000,110,000,52,000和40,000道尔顿的小成分。30S MSV RNA产生2个优势多肽，分子量分别为62,000和43,000道尔顿，次要成分分子量分别为72,000、40,000和18,000道尔顿。特异性抗血清免疫沉淀MuLV和MSV RNA产生的优势多肽是核心蛋白的前体。由MuLV RNA编码的180,000道尔顿分子量多肽通过抗血清对核心蛋白（p30）和逆转录酶进行免疫沉淀。因此，主要产品似乎是Pr65gag和Pr78gag；一个重要的小产品是Pr180gag-pol。两种系统合成的< 65,000道尔顿的产物大部分含有核心蛋白前体的序列，但MSV RNA产生的43,000和18,000道尔顿的MW多肽没有与抗血清沉淀到MULV蛋白。将纯化的酵母抑制物tRNA加入到35S MuLV RNA导向的翻译混合物中，Pr78gag的数量减少，而Pr180gag-pol的产量增加。琥珀色抑制tRNA的有效性约为赭色抑制tRNA和非抑制tRNA的3倍。这种抑制模式也见于[噬菌体]q . β合成酶基因中已建立的琥珀色突变（UAG）。(Q.beta。aml)，提示它是一个终止Pr78gag合成的UAG密码子。在MSV系统中，琥珀色抑制tRNA和赭色抑制tRNA在较小程度上显著增加了72,000道尔顿分子量多肽的合成，而62,000道尔顿蛋白的合成则略有减少。由于核心蛋白和逆转录酶基因之间在体内和体外都发生低水平的读通，并且在体外可以通过琥珀色抑制因子tRNA增强，这些结果表明，可能存在一种抑制机制控制了35S mRNA合成的核心蛋白和逆转录酶的相对数量。这种机制可能更广泛地应用于哺乳动物细胞的mRNA。
The virion RNA from Moloney murine leukemia virus (MuLV) and Moloney murine sarcoma virus (MSV) were translated in a micrococcal nuclease-treated [rabbit reticulocyte] cell-free system. The predominant polypeptides formed from 35S MuLV RNA were 78,000 and 65,000 daltons in MW, and minor components with MW of 180,000, 110,000, 52,000 and 40,000 daltons were also observed. The 30S MSV RNA yielded 2 predominant polypeptides of 62,000 and 43,000 daltons, and minor components about 72,000, 40,000 and 18,000 daltons in MW. The predominant polypeptides generated by both MuLV and MSV RNA were precursors of the core proteins by immunoprecipitation with specific antisera. The 180,000 dalton MW polypeptide encoded by MuLV RNA was immunoprecipitated both by antisera to the core protein (p30) and reverse transcriptase. The major products therefore appear to be Pr65gag and Pr78gag; an important minor product is Pr180gag-pol. Most of the products of < 65,000 daltons synthesized in either system contained sequences from the core protein precursor, but the 43,000 and 18,000 dalton MW polypeptides generated by MSV RNA did not precipitate with antisera to MULV proteins. When purified yeast suppressor tRNA was added to the translation mixture directed by 35S MuLV RNA, the amount of the Pr78gag was reduced, while the yield of the Pr180gag-pol was enhanced. Amber suppressor tRNA was about 3 times as effective as ochre suppressor tRNA and nonsuppressor tRNA. This pattern of suppression was also seen for an established amber mutation (UAG) in the synthetase gene of [phage] Q.beta. (Q.beta. aml), suggesting that it is a UAG codon which terminates synthesis of Pr78gag. In the MSV system, the amber suppressor tRNA, and to a lesser extent the ochre suppressor tRNA, markedly increased the synthesis of the 72,000 dalton MW polypeptide with a slight reduction of the 62,000 dalton protein. Since read-through between the core protein and reverse transcriptase genes occurs to a low level both in vivo and in vitro and can be enhanced in vitro by amber suppressor tRNA, these results suggest that a suppression mechanism may control the relative amounts of core protein and reverse transcriptase synthesized from 35S mRNA. Such a mechanism might be used more generally by mRNA from mammalian cells.