eIF2β is critical for eIF5-mediated GDP-dissociation inhibitor activity and translational control.
eIF2β is critical for eIF5-mediated GDP-dissociation inhibitor activity and translational control.
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DOI：
10.1093/nar/gkw657
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发表时间：
2016-11-16
影响因子：
14.9
通讯作者：
Pavitt GD
中科院分区：
文献类型：
--
作者：
Jennings MD;Kershaw CJ;White C;Hoyle D;Richardson JP;Costello JL;Donaldson IJ;Zhou Y;Pavitt GD
在蛋白质合成中，翻译因子eIF2将启动物tRNA与核糖体结合，促进启动密码子的选择。eIF2 GDP/GTP状态受eIF5 （GAP和GDI功能）和eIF2B （GEF和GDF活性）的调控，而eIF2α响应多种信号的磷酸化是翻译控制的主要点。在这里，我们描述了eIF2β中的生长抑制突变，该突变可以阻止eIF5 GDI并改变细胞对eIF2B活性降低的反应，包括GCN4翻译的控制。通过监测荧光核苷酸和启动物tRNA与纯化eIF2的结合，我们发现eIF2β突变不会影响这些配体对eIF2的内在亲和力，也不会干扰eIF2与43S起始前复合物组分的结合。相反，我们发现eIF2β突变阻止了eIF5 GDI稳定核苷酸与eIF2的结合，从而改变了eIF2•GDP/eIF5复合物的GDP脱出率。这使得细胞在eIF2B GEF活性降低的情况下生长，但在氨基酸饥饿反应中损害GCN4靶点的激活。这些发现为体内eIF5 GDI活性的重要性提供了支持，并证明eIF2β与eIF5协同作用，防止eIF2γ过早释放GDP，从而确保严格控制蛋白质合成起始。
In protein synthesis translation factor eIF2 binds initiator tRNA to ribosomes and facilitates start codon selection. eIF2 GDP/GTP status is regulated by eIF5 (GAP and GDI functions) and eIF2B (GEF and GDF activities), while eIF2α phosphorylation in response to diverse signals is a major point of translational control. Here we characterize a growth suppressor mutation in eIF2β that prevents eIF5 GDI and alters cellular responses to reduced eIF2B activity, including control of GCN4 translation. By monitoring the binding of fluorescent nucleotides and initiator tRNA to purified eIF2 we show that the eIF2β mutation does not affect intrinsic eIF2 affinities for these ligands, neither does it interfere with eIF2 binding to 43S pre-initiation complex components. Instead we show that the eIF2β mutation prevents eIF5 GDI stabilizing nucleotide binding to eIF2, thereby altering the off-rate of GDP from eIF2•GDP/eIF5 complexes. This enables cells to grow with reduced eIF2B GEF activity but impairs activation of GCN4 targets in response to amino acid starvation. These findings provide support for the importance of eIF5 GDI activity in vivo and demonstrate that eIF2β acts in concert with eIF5 to prevent premature release of GDP from eIF2γ and thereby ensure tight control of protein synthesis initiation.
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