CONTEXT EFFECTS ON N-6-ADENOSINE METHYLATION SITES IN PROLACTIN MESSENGER-RNA

The methylation of internal adenosine residues in mRNA only occurs wit hin GAC or AAC sequences. Although both of these sequence motifs are u tilized, a general preference has been noted for the extended sequence RGACU. Not all RGACU sequences in an mRNA are methylated and the mech anisms that govern the selection of methylation sites in mRNA remain u nclear. To address this problem we have examined the methylation of tr anscripts containing sequences of a natural mRNA, namely, bovine prola ctin mRNA. In this mRNA, a specific AGACU sequence in the 3' untransla ted region is the predominant site of methylation both in vivo and in vitro. The degree to which N-6-adenosine methyltransferase recognizes the sequence context of the consensus methylation site was explored by mutational analysis of the nucleotides adjacent to the core sequence as well as the extended regions in which the core element was found. O ur results indicate that efficient methylation depends on the extended five nucleotide consensus sequence but is strongly influenced by the context in which the consensus sequence occurs within the overall mRNA molecule. Furthermore, consensus methylation sites present in an RNA duplex are not recognized by the methyltransferase.