Yeast Trm7 interacts with distinct proteins for critical modifications of the tRNAPhe anticodon loop

Abstract

Post-transcriptional modification of the tRNA anticodon loop is critical for translation. Yeast Trm7 is required for 2′-O-methylation of C32 and N34 of tRNAPhe, tRNATrp, and tRNALeu(UAA) to form Cm32 and Nm34, and trm7-Δ mutants have severe growth and translation defects, but the reasons for these defects are not known. We show here that overproduction of tRNAPhe suppresses the growth defect of trm7-Δ mutants, suggesting that the crucial biological role of Trm7 is the modification of tRNAPhe. We also provide in vivo and in vitro evidence that Trm7 interacts with ORF YMR259c (now named Trm732) for 2′-O-methylation of C32, and with Rtt10 (named Trm734) for 2′-O-methylation of N34 of substrate tRNAs and provide evidence for a complex circuitry of anticodon loop modification of tRNAPhe, in which formation of Cm32 and Gm34 drives modification of m1G37 (1-methylguanosine) to yW (wyebutosine). Further genetic analysis shows that the slow growth of trm7-Δ mutants is due to the lack of both Cm32 and Nm34, and the accompanying loss of yW, because trm732-Δ trm734-Δ mutants phenocopy trm7-Δ mutants, whereas each single mutant is healthy; nonetheless, TRM732 and TRM734 each have distinct roles, since mutations in these genes have different genetic interactions with trm1-Δ mutants, which lack m2,2G26 in their tRNAs. We speculate that 2′-O-methylation of the anticodon loop may be important throughout eukaryotes because of the widespread conservation of Trm7, Trm732, and Trm734 proteins, and the corresponding modifications, and because the putative human TRM7 ortholog FTSJ1 is implicated in nonsyndromic X-linked mental retardation.