Cigan A M, Bushman J L, Boal T R, Hinnebusch A G
Section on Molecular Genetics of Lower Eukaryotes, National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD 20892.
Proc Natl Acad Sci U S A. 1993 Jun 1;90(11):5350-4. doi: 10.1073/pnas.90.11.5350.
In Saccharomyces cerevisiae, phosphorylation of the alpha subunit of translation initiation factor 2 (eIF-2) by protein kinase GCN2 stimulates translation of GCN4 mRNA. In mammalian cells, phosphorylation of eIF-2 alpha inhibits the activity of eIF-2B, the GDP-GTP exchange factor for eIF-2. We present biochemical evidence that five translational regulators of GCN4 encoded by GCD1, GCD2, GCD6, GCD7, and GCN3 are components of a protein complex that stably interacts with eIF-2 and represents the yeast equivalent of eIF-2B. In vitro, this complex catalyzes guanine nucleotide exchange on eIF-2 and overcomes the inhibitory effect of GDP on formation of eIF-2.GTP.Met-initiator tRNA(Met) ternary complexes. This finding suggests that mutations in GCD-encoded subunits of the complex derepress GCN4 translation because they mimic eIF-2 alpha phosphorylation in decreasing eIF-2B activity. Our results indicate that translational control of GCN4 involves a reduction in eIF-2B function, a mechanism used in mammalian cells to regulate total protein synthesis in response to stress.
在酿酒酵母中,蛋白激酶GCN2对翻译起始因子2(eIF-2)的α亚基进行磷酸化,可刺激GCN4 mRNA的翻译。在哺乳动物细胞中,eIF-2α 的磷酸化会抑制eIF-2B的活性,eIF-2B是eIF-2的GDP-GTP交换因子。我们提供了生化证据表明,由GCD1、GCD2、GCD6、GCD7和GCN3编码的GCN4的五个翻译调节因子是一种蛋白复合物的组成部分,该复合物与eIF-2稳定相互作用,相当于酵母中的eIF-2B。在体外,这种复合物催化eIF-2上鸟嘌呤核苷酸的交换,并克服GDP对eIF-2.GTP.Met起始tRNA(Met)三元复合物形成的抑制作用。这一发现表明,该复合物中由GCD编码的亚基发生突变会解除对GCN4翻译的抑制,因为它们在降低eIF-2B活性方面模拟了eIF-2α 的磷酸化。我们的结果表明,GCN4的翻译控制涉及eIF-2B功能的降低,这是哺乳动物细胞中用于响应应激调节总蛋白合成的一种机制。