A novel methyltransferase required for the formation of the hypermodified nucleoside wybutosine in eucaryotic tRNA.

We demonstrate that the product of the yeast open reading frame YML005w is required for wybutosine (yW) formation in the phenylalanine-accepting tRNA of the yeast Saccharomyces cerevisiae. tRNA isolated from a deletion mutant of the YML005w gene accumulates 4-demethylwyosine (ImG-14), a precursor lacking three of the methyl groups of the yW hypermodified base. Since the amino acid sequence of the YML005w gene contains the signature motifs of the seven beta-strand methyltransferases, we now designate the gene TRM12 for tRNA methyltransferase. Using pulse-chase labeling of intact yeast cells with S-adenosyl-L-[methyl-(3)H]methionine, we show that the methylesterified form of yW is metabolically stable.