Evidence That Phosphorylation of the -Subunit of eIF2 Does Not Essentially Inhibit mRNA Translation in Wheat Germ Cell-Free System.

A mechanism based on reversible phosphorylation of the -subunit of eukaryotic initiation factor 2 (eIF2) has been confirmed as an important regulatory pathway for the inhibition of protein synthesis in mammalian and yeast cells, while plants constitute the significant exception. We studied the induction of eIF2 phosphorylation in germinated wheat () embryos subjected to different adverse conditions. Data confirmed that formation of eIF2(P) was not a general response, as no phosphorylation was observed under salt, oxidative, or heat stress. Nevertheless, treatment by salicylic acid, UV-light, cold shock and histidinol did induce phosphorylation of eIF2 of wheat as has been established previously for eIF2 in Arabidopsis (). The influence of eIF2 phosphorylation on translation of reporter mRNA with different 5'-untranslated regions (5'UTRs) was studied in wheat germ cell-free system (WG-CFS), in which eIF2 was first phosphorylated either by heterologous recombinant human protein kinase, PKR (activated by double-stranded (ds)RNA), or by endogenous protein kinase GCN2 (activated by histidinol). Pretreatment of WG-CFS with PKR in the presence of dsRNA or with histidinol resulted in intense phosphorylation of eIF2; however, the translation levels of all tested mRNAs decreased by only 10-15% and remained relatively high. In addition, factor eIF2 from rabbit () bound GDP much more strongly than the homologous factor eIF2 from wheat germ. Furthermore, factor eIF2B was able to stimulate guanine nucleotide exchange (GDP→GTP) on eIF2 but had no effect on a similar exchange on eIF2. These results suggest that the mechanism of stress response eIF2 phosphorylation is not identical in all eukaryotes, and further research is required to find and study in detail new plant-specific mechanisms that may inhibit overall protein synthesis in plants under stress.