Effects of deprival of glucose or individual amino acids on polyribosome distribution and rate of protein synthesis in cultured mammalian cells

Abstract

Using Ehrlich ascites tumor cells in culture, we have examined the effects of omitting glucose or 13 essential amino acids, individually and in combination, on the rate of protein synthesis and the proportions of monomeric and polyribosomal ribosomes in the cells. These data have been used to determine the rate of peptide chain elongation, expressed as the synthesis rate per unit of polyribosomal ribosomes, and to infer a slowing of peptide chain initiation, relative to elongation, from an increase in the proportion of monomeric ribosomes. With omission of glucose from the culture medium, or addition of 2-deoxyglucose, the rates of peptide chain initiation and elongation decreased. These changes, which were rapidly reversed by glucose addition, occur in parallel with, and presumably are directly related to, a decrease in the intracellular ATP concentration. Omission of any one of 13 essential amino acids resulted in an immediate decrease of about 30% in the protein synthesis rate per cell, due largely to a decrease in initiation rate. Omitting combinations of two amino acids had about the same effect as the absence of all 13, producing an approx. 45% decrease in the synthesis rate per cell, again largely due to a slowing of initiation. Although amino acid restriction might be expected to exert a direct effect on the rate of both chain initiation and elongation through lowering of the intracellular aminoacyl-tRNA concentration, operation of some other rate-limiting mechanism is suggested by the fact that lack of any of the other 12 essential amino acids was as effective in slowing initiation as a lack of the initiating amino acid methionine. Evidence for such an indirect mechanism acting through limitation of the energy supply is the observation that deprival of amino acids resulted in a prompt and marked decrease in glucose utilization and in the amount of intracellular ATP. Thus, the effects of amino acid restriction on protein synthesis observed here may be mediated, at least in part, through a decrease in the rate of ATP generation.