title: Inhibition of rabbit reticulocyte lysate protein synthesis by heavy metal ions involves the phosphorylation of the alpha-subunit of the eukaryotic initiation factor 2.
abstract: The effect of heavy metal ions (in particular Cd2+, Hg2+, and Pb2+) on protein synthesis in hemin-supplemented reticulocyte lysates was investigated. Heavy metal ions were found to inhibit protein synthesis in hemin-supplemented lysates with biphasic kinetics. The shut off of protein synthesis occurred in conjunction with the phosphorylation of the alpha-subunit of the eukaryotic initiation factor (eIF) 2, the loss of reversing factor (RF) activity, and the disaggregation of polyribosomes. Addition of eIF-2 or RF to heavy metal ion-inhibited lysates restored protein synthesis to levels observed in hemin-supplemented controls. The stimulation of protein synthesis observed upon the addition of cAMP to heavy metal ion-inhibited lysates correlated with the inhibition of eIF-2 alpha phosphorylation and the restoration of RF activity. The partial restoration of protein synthesis observed upon the addition of MgGTP to heavy metal ion-inhibited lysates correlated with a partial inhibition of eIF-2 alpha phosphorylation. Addition of glucose 6-phosphate was found to have no effect on protein synthesis of eIF-2 alpha phosphorylation under these conditions. Antiserum raised to the reticulocyte heme-regulated eIF-2 alpha kinase inhibited the phosphorylation of eIF-2 alpha catalyzed by Hg2+-inhibited lysate. The inhibition of protein synthesis observed in the presence of heavy metal ions correlated with the relative biological toxicity of the ions. Highly toxic ions (AsO-2, Cd2+, Hg2+, Pb2+) inhibited protein synthesis by 50% at concentrations of 2.5-10 microM. Cu2+, Fe3+, and Zn2+, which are moderately to slightly toxic ions, inhibited protein synthesis by 50% at concentrations of 40, 250, and 300 microM, respectively. The data presented here indicate that heavy metal ions inhibit protein chain initiation in hemin-supplemented lysates by stimulating the phosphorylation of eIF-2 alpha apparently through the activation of the heme-regulated eIF-2 alpha kinase rather than through inhibition of the rate of eIF-2 alpha dephosphorylation.
authors: Department of Biochemistry, Oklahoma State University, Stillwater 74078-0454.
authors: R
authors: R
authors: Hurst
authors: J R
authors: JR
authors: Schatz
authors: R L
authors: RL
authors: Matts
mesh_terms: D000818:Animals; D002104:Cadmium / Q000494:pharmacology; D015852:Eukaryotic Initiation Factor-2; D006418:Heme / Q000494:pharmacology; D007700:Kinetics; D007854:Lead / Q000494:pharmacology; D046911:Macromolecular Substances; D008628:Mercury / Q000494:pharmacology; D008670:Metals / Q000494:pharmacology*; D010448:Peptide Initiation Factors / Q000378:metabolism*; D010766:Phosphorylation; D014176:Protein Biosynthesis / Q000187:drug effects*; D011506:Proteins / Q000378:metabolism*; D011817:Rabbits; D012156:Reticulocytes / Q000187:drug effects / Q000378:metabolism*
chemical_list: D015852:Eukaryotic Initiation Factor-2; D046911:Macromolecular Substances; D008670:Metals; D010448:Peptide Initiation Factors; D011506:Proteins; D002104:Cadmium; D007854:Lead; D006418:Heme; D008628:Mercury