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. 1974 Oct;71(10):3971–3975. doi: 10.1073/pnas.71.10.3971

Identification of Methylated Nucleosides in Messenger RNA from Novikoff Hepatoma Cells

Ronald Desrosiers 1, Karen Friderici 1, Fritz Rottman 1,*
PMCID: PMC434308  PMID: 4372599

Abstract

The poly(A) tract found in eukaryotic mRNA was used to study methylation in mRNA obtained from Novikoff hepatoma cells. Methyl labeling of RNA was achieved with L-[methyl-3H]methionine under conditions that suppress radioactive incorporation into the purine ring. RNA that contains a poly(A) segment was obtained from polysomal RNA by chromatography on oligo(dT)-cellulose. Sucrose density gradient centrifugation of this RNA revealed a pattern expected for mRNA. The composition of the methyl-labeled nucleosides in the RNA was analyzed after complete enzymatic degradation to nucleosides. By use of DEAE-cellulose (borate) chromatography, which separates 2′-O-methylnucleosides from normal and base-methylated nucleosides, about 50% of the radioactivity was recovered in the 2′-O-methylnucleoside fraction and 50% in the base-methylnucleoside fraction. High-speed liquid chromatography (Aminex A-5) of the 2′-O-methylnucleoside fraction produced four peaks coincident with the four 2′-O-methylnucleoside standards. Analysis of the base-methylnucleoside fraction revealed a unique pattern. While ribosomal RNA and tRNA possessed complex base-methylnucleoside patterns, the distribution in mRNA was quite simple, consisting predominantly of N6-methyladenosine. These results demonstrate a unique distribution of methylated nucleosides in mRNA. By analogy to ribosomal RNA synthesis, the presence of methylnucleosides in mRNA may reflect a cellular mechanism for the selective processing of certain mRNA sequences.

Keywords: RNA methylation, RNA processing, methylnucleoside composition

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

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