Eukaryotic translation initiation factor 3 (eIF3) and eIF2 can promote mRNA binding to 40S subunits independently of eIF4G in yeast
Eukaryotic translation initiation factor 3 (eIF3) and eIF2 can promote mRNA binding to 40S subunits independently of eIF4G in yeast
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DOI：
10.1128/mcb.26.4.1355-1372.2006
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发表时间：
2006-02-01
影响因子：
5.3
通讯作者：
Nielsen, KH
中科院分区：
文献类型：
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作者：
Jivotovskaya, AV;Valásek, L;Nielsen, KH
体外多种起始因子（包括 eIF3、eIF1、eIF5 和 eIF1A）会刺激真核翻译起始因子 2 (eIF2)-GTP-Met-tRNA(i)(Met) 三元复合物向 40S 核糖体的募集。 mRNA 的募集被认为需要 eIF4F 和 eIF3 的功能，后者充当核糖体和 eIF4F 4G 亚基之间的接头。为了确定体内这些反应的因子要求，我们检查了酿酒酵母细胞中去除 eIF2、eIF3、eIF5 或 eIF4G 对三元复合物、其他起始因子和 RPL41A mRNA 与天然 43S 和 48S 预起始复合物结合的影响。耗尽 eIF2、eIF3 或 eIF5 会减少由这三个因子和 eIF1 组成的多因子复合物 (MFC) 所有成分的 40S 结合，支持 MFC 成分耦合 40S 结合的机制。 40S 结合的 mRNA 在 eIF5 耗尽的细胞中强烈积累，尽管 MFC 与 40S 亚基的结合因 eIF5 耗尽而减少。因此，刺激三元复合物的 GTP 酶活性（60S 亚基体外连接的先决条件）可能是体内 eIF5 的限速功能。耗尽 eIF2 或 eIF3 会损害 mRNA 与游离 40S 亚基的结合，但耗尽 eIF4G 会意外地导致 mRNA 在 40S 亚基上积累。因此，对于至少一些mRNA与天然预起始复合物的稳定结合，似乎比eIF4G更关键地需要eIF3和eIF2，并且eIF4G在体内48S复合物组装的下游步骤具有限速功能。
Recruitment of the eukaryotic translation initiation factor 2 (eIF2)-GTP-Met-tRNA(i)(Met) ternary complex to the 40S ribosome is stimulated by multiple initiation factors in vitro, including eIF3, eIF1, eIF5, and eIF1A. Recruitment of mRNA is thought to require the functions of eIF4F and eIF3, with the latter serving as an adaptor between the ribosome and the 4G subunit of eIF4F. To define the factor requirements for these reactions in vivo, we examined the effects of depleting eIF2, eIF3, eIF5, or eIF4G in Saccharomyces cerevisiae cells on binding of the ternary complex, other initiation factors, and RPL41A mRNA to native 43S and 48S preinitiation complexes. Depleting eIF2, eIF3, or eIF5 reduced 40S binding of all constituents of the multifactor complex (MFC), comprised of these three factors and eIF1, supporting a mechanism of coupled 40S binding by MFC components. 40S-bound mRNA strongly accumulated in eIF5-depleted cells, even though MFC binding to 40S subunits was reduced by eIF5 depletion. Hence, stimulation of the GTPase activity of the ternary complex, a prerequisite for 60S subunit joining in vitro, is likely the rate-limiting function of eIF5 in vivo. Depleting eIF2 or eIF3 impaired mRNA binding to free 40S subunits, but depleting eIF4G led unexpectedly to accumulation of mRNA on 40S subunits. Thus, it appears that eIF3 and eIF2 are more critically required than eIF4G for stable binding of at least some mRNAs to native preinitiation complexes and that eIF4G has a rate-limiting function at a step downstream of 48S complex assembly in vivo.