Biochemical studies on the mechanism of action of liver poisons I. Inhibition of protein synthesis

Administration of ethionine to rats was followed by a rapid decline in the uptake of labeled amino acids into the liver proteins. The magnitude of the decrease in the extent of the protein labeling remained unaltered when different radioactive amino acids were used as tracers, or when the amount of the amino acid injected was increased from a tracer dose up to a level in excess of the endogenous pool size. It was inferred that the reduced amino acid incorporation in vivo was not attributable to isotope dilution by a possible elevation of the free amino acid pools, but reflected an inhibition of protein synthesis. In rats treated with ethionine the derangement of protein synthesis in vivo exhibited an earlier onset and a faster time course than the decay of the cell-free, endogenously coded amino acid incorporation coinciding with an enhancement of the polyuridylic acid-directed phenylalanine polymerization and a disaggregation of the polysomes. This discrepancy between the inhibitory effects of ethionine observed in the intact animals and in the cell-free preparations was particularly prominent in male rats in which the dissociation of the polysomes and the deterioration of the cell-free protein-synthesizing capacity were much less pronounced than in the female rats. In contrast, the extent of hepatic ATP depletion was equally severe in both sexes. Injection of adenine or its nucleotide congeners together with ethionine into female rats prevented the fall of the liver ATP level as well as the decay of the protein-synthesizing capacity as measured in vitro, but failed to relieve appreciably the inhibition of protein synthesis in vivo. Paralell examination of the polysome patterns revealed that the exogenously supplied ATP precursors and ethionine induced a synergistic inhibition of protein synthesis in vivo, different from the injury caused by ethionine alone. This inhibition, characterized by the preservation of the intact polysomal structure, suggested an interference with the completion of the nascent peptide chains as the underlying mechanism of the phenomenon concerned. A similar type of inhibition was obtained by simultaneous administration of adenine mucleotides and an excess of methionine or other amino acids. The overall data are consistent with the interpretation that the synergistic block of peptide chain elongation "freezes" the polysomal assembly and thereby prevents the expression of the initiation lesion inherent in the deleterious effect of ethionine on the protein-synthesizing system. This explanation is borne out by the observation that pretreatment with a mixture consisting of 3′(2′)-AMP and either ethionine or methionine obviated the carbon tetrachloride-induced disaggregation of the polysomes and the concomitant decline of the cell-free amino acid-incorporating activity. The possible mechanisms underlying the noxiuos effects of ethionine on protein synthesis are discussed with special reference to the crucial role of ATP depletion in the pathogenesis of ethionine poisoning.