Inhibition of methylation at two internal N6-methyladenosine sites caused by GAC to GAU mutations

• Published in: The Journal of biological chemistry Vol. 262; no. 7; pp. 3422 - 3427
• Main Authors: Kane, S.E., Beemon, K.
• Format: Journal Article
• Language: English
• Published: Bethesda, MD Elsevier Inc 05.03.1987; American Society for Biochemistry and Molecular Biology
• Subjects: Adenosine - analogs & derivatives; Adenosine - metabolism; Analytical, structural and metabolic biochemistry; Animals; Avian Sarcoma Viruses - genetics; Base Sequence; Biological and medical sciences; Chick Embryo; DNA, Recombinant; Fundamental and applied biological sciences. Psychology; Methylation; Mutation; Nucleic Acid Hybridization; Nucleic acids; RNA, Messenger - genetics; Rna, ribonucleoproteins; RNA, Viral - genetics; RNA, Viral - metabolism; Transfection; Virus; Oncovirinae; RNA; Oncogenic virus; Type C oncovirus; Retroviridae; Rous sarcoma virus; Inhibition; Mutation; Methylation
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1016/S0021-9258(18)61520-0

We previously have mapped N6-methyladenosine (m6A) sites within the genomic RNA of Rous sarcoma virus (RSV). The results of that study and of experiments using inhibitors of methylation suggest that m6A might be involved in mRNA processing events. We describe an approach for directly analyzing the function of m6A in RNA and for studying the sequence specificity of the m6A methylase. Two sites of methylation in RSV (nucleotides 7414 and 7424) were altered by oligonucleotide-directed mutagenesis. The highly conserved GAC consensus sequence at those sites was changed to GAU. The new sequences were no longer methylated in the RSV genomic RNA; the GAC sequence was required for efficient base modification at those two adenosines. The altered m6A pattern did not affect viral RNA processing or the viral life cycle within infected cells.