Ultrastructural localization of phosphorylated eIF2α [eIF2α(P)] in rat dorsal hippocampus during reperfusion

During post-ischemic brain reperfusion there is a substantial reduction of protein synthesis in selectively vulnerable neurons. Normal protein synthesis requires a functional translation initiation complex, a key element of which is eukaryotic initiation factor 2 (eIF2), which in a complex with GTP introduces the met-tRNA(i). Phosphorylation of Ser(51) on the alpha subunit of eIF2 [eIF2 alpha(P)] generates a competitive inhibitor of eIF2B, thereby preventing the replenishment of GTP onto eIF2, thus blocking translation initiation. It has been shown that the conditional expression of an eIF2 alpha mutant (Asp substituted for Ser51) imitating the negative charge of Ser(51) (P) induces apoptosis. During the first 10 min of post-ischemic reperfusion, there is an approximately 20-fold increase in eIF2 alpha(P) seen in the cytoplasm of CA1 hippocampal neurons, and, by 1 h, there is also accumulation of eIF2 alpha(P) in the nucleus. We utilized post-embedding electron microscopical immunogold methods to examine the localization of eIF2 alpha(P) during reperfusion. Immunogold particles (10 nm) were concentrated chiefly along the rough endoplasmic reticulum and in association with the membranes of the nuclear envelope in CA1 neurons. Aggregations of gold particles in the nucleus were concentrated: (1) within and around the nucleolus, (2) associated to strands of heterochromatin, and (3) along putative nuclear filaments. The presence of eIF2 alpha(P) in the nucleolus probably reflects its association with nascent ribosomal subunits. The P-subunit of eIF2 has a zinc finger and polylysine blocks analogous to these on other proteins that affect transcription. The association of eIF2 alpha(P) with chromatin may have important implications for transcription.