Methylation of ribonucleic acid in a cell-free system from mouse myeloma cells.

Abstract

Isolated nuclei incorporate few methyl groups into RNA when they are incubated with S-adenosyl[methyl-3H3]methionine and four ribotriphosphates. When the nuclei were supplemented with a soluble total cell protein extract, the incorporation of methyl groups into RNA was stimulated 3-6-fold. All classes of RNA were methylated. Methylation of the 2'-OH of ribose and the bases of ribosomal RNA occurred predominantly on endogenous ribosomal RNA precursors, with a minority (20%) occurring on the newly synthesized rRNA precursor. Methylation of the tRNA precursor occurred on both endogenous (40%) and newly synthesized (60%) molecules. The methylation of adenosine in hnRNA occurred predominantly on molecules transcribed in vitro and was sensitive to 1 microgram/mL alpha-amanitin. A final site of methylation was the 7 position of guanosine of the cap structure. About 10% of the RNA polymerase II transcripts were capped in vitro. Capping was blocked 90% by 1 microgram/mL alpha-amanitin and was independent of the presence of the cell protein extract.