Studies on the mechanism of the inhibition of protein synthesis induced by intracellular ATP depletion

Abstract

The deleterious effect of intracellular ATP depletion on the protein-synthesizing system was studied in rabbit reticulocytes and Landschütz ascites tumor cells. The progressive decline of the ATP content of the cells induced by anaerobic incubation in glucose-free Krebs-Ringer salt solution was attended by a prompt inhibition of protein synthesis. The onset of the arrest of protein synthesis in the course of incubation under nitrogen, coincident with a 20–30 % decrease of the ATP level, entailed no impairment of the cell-free amino acid-incorporating activity and was readily reversible by reaeration of the cells. This early inhibition showed no correlation with the degree of ATP deprivation but appeared to be determined by the inhibitory effect of the accumulating ADP and AMP on the process of peptide chain elongation. Extensive ATP depletion following prolonged anaerobic incubation gave rise to a lesion characterized by dissociation of the polysomes and a concomitant decline of the cell-free amino acid-incorporating activity. However, the expression of this lesion was masked by factors interfering with the peptide chain elongation. Therefore, drastic dissociation of the polysomes within the cells occurred only during the brief anaerobic-aerobic transition period following readmission of air when the block of the chain elongation by the products of ATP splitting had been relieved by partial resynthesis of ATP. On prolonging the aerobic reincubation of the cells, progressive reversal of the derangement of protein synthesis took place. It was concluded that the lesion concerned was localized at the stage of peptide initiation. The initiation defect caused by severe ATP deficiency was shown to reside exclusively in the cell-sap fraction and to differ essentially from the ribosomal lesion induced by NaF. In addition, the detrimental effect of NaF on protein synthesis proved to be entirely unrelated to the inhibitory action of this poison on the energy metabolism. An inhibitory pattern essentially similar to the one above was obtained when ATP depletion was brought about by incubating the cells in the presence of 2,4-dinitrophenol. Furthermore, the alterations induced by the uncoupling action of dinitrophenol on the oxidative phosphorylation were completely reversible by washing away the uncoupling agent. The mechanism underlying the above alterations of the protein-synthesizing system are discussed in light of the pertinent data, and the possible regulatory implication of the inhibitory influence of the products of ATP breakdown on peptide chain elongation is considered.