Identification of the yeast gene encoding the tRNA m¹G methyltransferase responsible for modification at position 9

Abstract

Methylation of tRNA at the N-1 position of guanosine to form m¹G occurs widely in nature. It occurs at position 37 in tRNAs from all three kingdoms, and the methyltransferase that catalyzes this reaction is known from previous work of others to be critically important for cell growth in Escherichia coli and the yeast Saccharomyces cerevisiae. m¹G is also widely found at position 9 in eukaryotic tRNAs, but the corresponding methyltransferase was unknown. We have used a biochemical genomics approach with a collection of purified yeast GST-ORF fusion proteins to show that m¹G₉ formation of yeast tRNA^Gly is associated with ORF YOL093w, named TRM10. Extracts lacking Trm10p have undetectable levels of m¹G₉ methyltransferase activity but retain normal m¹G₃₇ methyltransferase activity. Yeast Trm10p purified from E. coli quantitatively modifies the G₉ position of tRNA^Gly in an S-adenosylmethionine-dependent fashion. Trm10p is responsible in vivo for most if not all m¹G₉ modification of tRNAs, based on two results: tRNA^Gly purified from a trm10-Δ/trm10-Δ strain is lacking detectable m¹G; and a primer extension block occurring at m¹G₉ is removed in trm10-Δ/trm10-Δ-derived tRNAs for all 9 m¹G₉-containing species that were testable by this method. There is no obvious growth defect of trm10-Δ/trm10-Δ strains. Trm10p bears no detectable resemblance to the yeast m¹G₃₇ methyltransferase, Trm5p, or its orthologs. Trm10p homologs are found widely in eukaryotes and many archaea, with multiple homologs in several metazoans, including at least three in humans.

Keywords

• Saccharomyces cerevisiae
• S-adenosylmethionine
• TRM10
• YOL093w
• 1-methylguanosine