Evidence That Ternary Complex (eIF2-GTP-tRNA_i^Met)–Deficient Preinitiation Complexes Are Core Constituents of Mammalian Stress Granules

TLDR

Phosphorylation of eIF2α during environmental stress blocks translation by depleting the ternary complex, causing untranslated mRNAs to be routed to stress granules that depend on TIA‑1/TIAR and whose relationship with polysomes remains unclear. The study proposes a translational checkpoint model in which TIA proteins and eIF2 act as functional antagonists, and the absence of ternary complex drives stress granule assembly. In arsenite‑stressed cells, most 48S preinitiation complex components (small ribosomal subunits, eIF3, eIF4E, eIF4G) are recruited to stress granules, whereas eIF2 is excluded; a phosphomimetic eIF2α mutant forms SGs of comparable composition, and phospho‑eIF2α is also recruited to disassembling SGs during recovery.

Abstract

Environmental stress-induced phosphorylation of eIF2α inhibits protein translation by reducing the availability of eIF2-GTP-tRNA i Met, the ternary complex that joins initiator tRNA Met to the 43S preinitiation complex. The resulting untranslated mRNA is dynamically routed to discrete cytoplasmic foci known as stress granules (SGs), a process requiring the related RNA-binding proteins TIA-1 and TIAR. SGs appear to be in equilibrium with polysomes, but the nature of this relationship is obscure. We now show that most components of the 48S preinitiation complex (i.e., small, but not large, ribosomal subunits, eIF3, eIF4E, eIF4G) are coordinately recruited to SGs in arsenite-stressed cells. In contrast, eIF2 is not a component of newly assembled SGs. Cells expressing a phosphomimetic mutant (S51D) of eIF2α assemble SGs of similar composition, confirming that the recruitment of these factors is a direct consequence of blocked translational initiation and not due to other effects of arsenite. Surprisingly, phospho-eIF2α is recruited to SGs that are disassembling in cells recovering from arsenite-induced stress. We discuss these results in the context of a translational checkpoint model wherein TIA and eIF2 are functional antagonists of translational initiation, and in which lack of ternary complex drives SG assembly.