Identification of Methylated Nucleosides in Messenger RNA from Novikoff Hepatoma Cells

来自 Semantic Scholar

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阅读量：

641

作者：

R Desrosiers，K Friderici，F Rottman

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摘要：

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-labeld 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.

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关键词：

Biochemistry RNA methylation RNA processing Methylnucleoside composition

DOI：

10.1073/pnas.71.10.3971

被引量：

536

年份：

1974