The role of amino acids in the regulation of protein synthesis in perfused rat liver. II. Effects of amino acid deficiency on peptide chain initiation, polysomal aggregation, and distribution of albumin mRNA.
Decreased rates of protein synthesis which occurred in rat livers perfused with amino acid-deficient medium were accompanied by a loss of polysomes and a doubling of concentrations of ribosomal subunits and monomers as compared to unperfused liver or livers perfused with amino acid-supplemented medium. The loss of polysomes was not the result of mRNA degradation because this effect could be reversed by addition of amino acids to the perfusion medium. Instead, loss of polysomes indicated an impairment in peptide chain initiation. To determine whether the block of initiation could be the result of sequestration of mRNA in untranslatable pools, the content of albumin mRNA in membrane-bound and free polysomes and in the non-polysomal fraction isolated on sucrose density gradients was determined by hybridization to an albumin cDNA. In livers perfused with an amino acid-supplemented medium, 90% of the albumin mRNA was found in bound polysomes, and with amino acid-deficient medium, this value was decreased to 80%. Thus, there was no indication of a significant pool of mRNA which was not being translated. The block of initiation was, however, accompanied by a marked change in density and methionine-binding characteristics of the 40 S ribosomal subunit in livers perfused with deficient medium as compared to livers perfused with supplemented medium or to unperfused livers. There was a relative loss of 40 S subunits in a low density form, 1.41 g/cm3, an increase in the proportion in a high density form, 1.48 g/cm3, and a decrease in binding of [35S]methionine to 40 S ribosomal subunits. These changes provide evidence of reduced rates of formation of the 40 S initiation complex.
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000378:metabolism*; D000818:Animals; D001483:Base Sequence; D008099:Liver / Q000378:metabolism*; D008297:Male; D010442:Peptide Chain Initiation, Translational; D010477:Perfusion; D011132:Polyribosomes / Q000378:metabolism*; D011506:Proteins / Q000235:genetics*; D012333:RNA, Messenger / Q000235:genetics*; D051381:Rats; D011919:Rats, Inbred Strains; D012709:Serum Albumin / Q000235:genetics*
D000596:Amino Acids; D011506:Proteins; D012333:RNA, Messenger; D012709:Serum Albumin