CHANGES IN eIF2α PHOSPHORYLATION IN RESPONSE TO NUTRIENT DEFICIENCY AND OTHER STRESSORS IN FISH
Place, Allen R
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The present study investigates the response of two teleost species to stressors as measured by phosphorylation of the α-subunit of the translational initiation factor, eIF2. The phosphorylation of the translational initiation factor, eIF2, on its α-subunit is an adaptive response to a variety of stressors in eukaryotes from protists to vertebrates. There are four eIF2α-specific kinases in most vertebrates, GCN2, PERK, PKR and HRI, each of which can be activated by different stressors. Two of these eIF2α-kinases, GCN2 and PERK, respond to changes in nutritional status. It was of particular interest to determine whether eIF2α phosphorylation can be used as an early marker to evaluate diets in fish. However, eIF2α phosphorylation could also be of use in monitoring other stressors of fish in aquaculture. The increase in global aquaculture production of fish has increased interest in the optimization of fish diets and health to increase production and sustainability. Studies were initiated in a zebrafish Danio rerio cell line, ZFL cells, to lay the groundwork for looking at eIF2α phosphorylation in fish and in species of more interest to aquaculture. All the eIF2α-kinases are present in the zebrafish genome and are expressed in ZFL cells. Two forms of eIF2α are expressed in ZFL cells, eIF2α-a and eIF2α-b, with eIF2α-b transcripts ~5-fold higher than those of eIF2α-a. The two gene products are 96 % identical at the amino acid level and are identical at the phosphorylation and kinase docking sites. Phosphorylation of eIF2α in ZFL cells is increased by a variety of agents/conditions; starvation, leucinol, endoplasmic reticulum stress, poly(I)poly(C) and N-methylprotoporphyrin, consistent with activation of the eIF2α kinases GCN2, PERK, PKR and HRI, respectively. Application of the same analyses to a new cell line from a marine fish, cobia Rachycentron canadum, shows that these cells are also responsive to activators of GCN2 and PERK. Cloning of eIF2α cDNA from cobia has shown close identity to the zebrafish eIF2αs. Although cobia has two eIF2α transcripts, the coding sequence of each is identical. Preliminary studies have shown that in cobia juveniles, diet, probiotics and water temperature all affect the phosphorylation state of eIF2α.