TRANSLATION INITIATION FACTOR-4A FROM SACCHAROMYCES-CEREVISIAE - ANALYSIS OF RESIDUES CONSERVED IN THE D-E-A-D FAMILY OF RNA HELICASES
TRANSLATION INITIATION FACTOR-4A FROM SACCHAROMYCES-CEREVISIAE - ANALYSIS OF RESIDUES CONSERVED IN THE D-E-A-D FAMILY OF RNA HELICASES
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DOI：
10.1128/mcb.11.7.3463
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发表时间：
1991-07-01
影响因子：
5.3
通讯作者：
LINDER, P
中科院分区：
文献类型：
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作者：
SCHMID, SR;LINDER, P
真核生物翻译起始因子4A （eIF-4A）具有体外解旋酶活性，允许双链RNA解绕。这种活性依赖于ATP水解和另一种翻译起始因子eIF-4B的存在。这两个起始因子被认为解开mRNA二级结构，为核糖体结合和翻译起始做准备。为了进一步表征eIF-4A在细胞翻译中的功能及其与翻译机制中其他元件的相互作用，我们分离了酿酒酵母编码eIF-4A的TIF1和TIF2基因的突变。我们发现D-E-A-D蛋白家族的三个高度保守结构域，编码eIF-4A和其他RNA解旋酶，对蛋白质功能至关重要。只有在极少数情况下，我们才能在不影响细胞生长的情况下进行保守替代。突变体的生长与体内翻译率之间存在明显的相关性。一种导致温度敏感表型的突变表明，在转向不允许的温度后，翻译活性立即下降。这些体内结果证实了先前的体外数据，表明翻译绝对依赖于TIF1和TIF2基因产物。
The eukaryotic translation initiation factor 4A (eIF-4A) possesses an in vitro helicase activity that allows the unwinding of double-stranded RNA. This activity is dependent on ATP hydrolysis and the presence of another translation initiation factor, eIF-4B. These two initiation factors are thought to unwind mRNA secondary structures in preparation for ribosome binding and initiation of translation. To further characterize the function of eIF-4A in cellular translation and its interaction with other elements of the translation machinery, we have isolated mutations in the TIF1 and TIF2 genes encoding eIF-4A in Saccharomyces cerevisiae. We show that three highly conserved domains of the D-E-A-D protein family, encoding eIF-4A and other RNA helicases, are essential for protein function. Only in rare cases could we make a conservative substitution without affecting cell growth. The mutants show a clear correlation between their growth and in vivo translation rates. One mutation that results in a temperature-sensitive phenotype reveals an immediate decrease in translation activity following a shift to the nonpermissive temperature. These in vivo results confirm previous in vitro data demonstrating an absolute dependence of translation on the TIF1 and TIF2 gene products.