Factors involved in increased protein synthesis in rat kidney after administration of DDT

Homogenates of kidney cortex obtained from control rats and rats treated with DDT have been separated into microsomes or ribosomes, and into postmicrosomal (S105) supernatant fraction or pH 5 supernatant fraction. The incorporation of [¹⁴C]phenylalanyl-tRNA into peptide was increased when microsomes derived from kidneys of DDT-treated rats were incubated with pH 5 supernatant fraction from control rats. Elongation factors (EF) 1 and 2, necessary for the binding of aminoacyltRNA to ribosomes and for translocation of peptidyl-tRNA from the A site to the P site of ribosomes, were present in the pH 5 supernatant fractions of kidney of control and DDT-treated rats and these fractions were incubated with KCl-washed ribosomes obtained from livers of control rats. The results provided evidence that the increased incorporation observed with the pH 5 supernatant fraction obtained from the DDT-treated animals could not be attributed to decreased ribonuclease activity or to increased elongation factor 2 activity but was due to an increase in elongation factor l activity.     

Protein synthesis catalyzed by thiol blocked ribosome preparations

Ribosome preparations respond heterogeneously to thiol blocking agents such as N-ethylmaleimide. About 55% of the particles rapidly lose the ability to catalyze peptide bond formation. The remaining 45% are immune. The only function related to peptide bond formation that appears abnormal in inactivated ribosomes is messenger RNA-directed transfer RNA binding. The change, however, is small relative to the alteration in activity observed. Thus it appears that thiol blocking, which selectively damages 30 S subunits, renders about half the 30 S subunits unable to co-ordinate properly with 50 S subunits in peptide bond formation. The implications of these results with regard to ribosome heterogeneity are discussed.     

Interactions between mutations affecting ribosome synthesis in Escherichia coli

RNA synthesis was followed during amino acid starvation of strains of Escherichia coli that contained both the relaxed (relA) mutation and a mutation affecting ribosome assembly that results in oversynthesis of RNA. The ribosome mutation did not by itself lead to relaxedness. The relaxed mutation could be expressed in organisms that contained the ribosome mutation.     

Temperature sensitive mutations affecting ribosome synthesis in Saccharomyces cerevisiae

Studies have been carried out on the effects of mutations in nine distinct genes which regulate ribosome biosynthesis in yeast. Although some mutants have more extensive effects than others, in each case there is an inhibition by 50 to 80% of the synthesis of ribosomal precursor RNA. In each case there is an inhibition of the processing of that RNA which is made, ranging from 80 to 95%. In each case there is degradation of 50 to 70% of that RNA which is processed. Much of the RNA which survives to become the mature 25 s and 18 s species is found in functioning polyribosomes.     

Factors affecting protein synthesis during biotin deficiency inAspergillus nidulans

Biotin deficiency in Aspergillus nidulans resulted in a 70% increase of the protein content and increased levels of free and bound aspartate, glutamate, serine, leucine and methionine. Likewise, the activities of NADP+ glutamate dehydrogenase, NAD+ gluatmate dehydrogenase, aspartate aminotransferase and alanine aminotransferase were significantly increased. The total RNA content increased while the DNA content was unaffected. The rRNA/tRNA ratio remained higher in biotin-deficient cells. Supplementation of glutamate, aspartate, serine, leucine and methionine to the culture medium raised the rRNA/tRNA ratio, and the difference observed in the qualitative and the quantitative patterns of protein and dry cell mass between normal and biotin-deficient cultures was abolished.     

Fractionation of the ribosome inactivating protein preparations with triazine dyes

Aspergillins are ribosome-inactivating proteins (RIPs), isolated from several strains of Aspergillus. The interaction between Cibacron Blue F3GA and two members of this family, alpha sarcin and mitogillin, and other RIPs of type I, was studied. Alpha sarcin retention depended on pH and ionic strength. By chromatography on Affi-Gel Blue in mild experimental conditions, mitogillin and PAP-I did not interact with the dye, whereas 40% of alpha sarcin and 70-90% of briodin, RTA and gelonin were recovered in the bound fraction. In all cases, the major fraction showed a higher toxicity level in protein synthesis inhibition assays. The unbound alpha sarcin, conjugated with the anti-ovarian carcinoma monoclonal antibody MOv17, showed on OVCA 432 a cytotoxicity which was 900 times higher than that exerted by the alpha sarcin alone.     

A paradigm for ribosome fidelity modulation in protein synthesis

The ribosome, an integral component of the protein synthetic apparatus, plays a role in mediating the fidelity of the translation of genetic information. It is proposed here that ribosome fidelity modulation can be achieved in part through ribosomal RNA interactions with messenger RNA. These nucleic acid interactions generate alterations in the configuration of the ribosome which enhance the particles affinity for classes on specific transfer RNAs. These changes are directly related to the information content of a given messenger RNA which is being translated and yield an increase in the fidelity of translation. The molecular mechanism for effecting the ribosomal structural changes in response to the genetic code is described and correlated with the properties of ribosomal proteins and transfer RNAs.