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Methylated nucleotides block 5' terminus of HeLa cell messenger RNA

C M Wei, A Gershowitz, B Moss

    Cell
    |April 1, 1975
    PubMed

    Insights

    HeLa cell messenger RNA (mRNA) contains methylated bases, with 6-methyladenosine being the most abundant. A unique 7-methylguanosine cap structure at the 5' terminus of mRNA may be crucial for eukaryotic cell function.

    Area of Science:

    • Molecular Biology
    • Biochemistry
    • Eukaryotic Gene Expression

    Background:

    • Polyadenylylated [poly(A)+] mRNA plays a critical role in gene expression.
    • Understanding mRNA modifications, such as methylation, is essential for deciphering gene regulation.

    Purpose of the Study:

    • To investigate the types and locations of methylated nucleosides in poly(A)+ mRNA from HeLa cells.
    • To characterize the 5' terminal structure of eukaryotic mRNA.

    Main Methods:

    • Isolation of poly(A)+ RNA using poly(U)-Sepharose chromatography.
    • Quantification of methylation levels using radiolabeling ([3H-methyl]-methionine and 14C-uridine) and 3H/14C ratios.
    • Identification of methylated nucleosides via chromatographic and chemical analyses.

    Main Results:

    • Poly(A)+ RNA contains approximately two methyl groups per 1000 nucleotides.
    • Identified four 2'-O-methylribonucleosides and two base-methylated derivatives: 6-methyladenosine (6MeA) and 7-methylguanosine (7MeG).
    • 6MeA constituted ~50% of methylated ribonucleosides; 7MeG (~10%) was exclusively at the 5' terminus, forming a cap structure (7MeGpppNmepNmep-).

    Conclusions:

    • Eukaryotic mRNA possesses a methylated 5' cap structure, primarily 7-methylguanosine.
    • This blocked 5' terminus is likely a characteristic feature of functional eukaryotic mRNAs.
    • The findings suggest conserved mechanisms for mRNA processing and translation initiation across eukaryotes.

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