Methylation of newly synthesized ribonucleic acid by isolated rat liver nuclei. Characterization of the ribonucleic acid synthesized by nuclei from starved animals

Abstract

1. Nuclei from rat liver incubated with S-adenosyl[methyl-14C]methionine incorporated radioactivity into RNA and into lipid and protein. 2. All of the labelled RNA was extracted from the nuclei with trichloroacetic acid at 90°C. 3. The [14C]methyl-group incorporation into the hot-trichloroacetic acid extract was 30% inhibited by the addition of actinomycin D (100μg/mg of DNA) or by the omission of CTP, GTP and UTP. 4. Assuming that the main substrate for this triphosphate-dependent methylation was newly synthesized precursor rRNA containing one methyl group/30 uridylate residues, it was calculated that approx. 60% of the [14C]UMP incorporated under similar conditions represented precursor rRNA synthesis. 5. In agreement with this, low concentrations of actinomycin D (approx. 1μg/mg of DNA) sufficient to abolish the triphosphate-dependent incorporation of [14C]methyl group inhibited 68% of the [14C]UMP incorporation. 6. The incorporation of [14C]UMP by nuclei from starved animals decreased progressively with increasing periods of starvation, whereas the triphosphate-dependent [14C]methyl-group incorporation was not further decreased after 1 day of starvation. 7. This suggests that precursor rRNA synthesis decreased within 1 day whereas other species of RNA were affected only after longer periods of starvation.