Conservation of an intricate circuit for crucial modifications of the tRNAPhe anticodon loop in eukaryotes

Abstract

Post-transcriptional tRNA modifications are critical for efficient and accurate translation, and have multiple different roles. Lack of modifications often leads to different biological consequences in different organisms, and in humans is frequently associated with neurological disorders. We investigate here the conservation of a unique circuitry for anticodon loop modification required for healthy growth in the yeast Saccharomyces cerevisiae. S. cerevisiae Trm7 interacts separately with Trm732 and Trm734 to 2′-O-methylate three substrate tRNAs at anticodon loop residues C32 and N34, and these modifications are required for efficient wybutosine formation at m1G37 of tRNAPhe. Moreover, trm7Δ and trm732Δ trm734Δ mutants grow poorly due to lack of functional tRNAPhe. It is unknown if this circuitry is conserved and important for tRNAPhe modification in other eukaryotes, but a likely human TRM7 ortholog is implicated in nonsyndromic X-linked intellectual disability. We find that the distantly related yeast Schizosaccharomyces pombe has retained this circuitry for anticodon loop modification, that S. pombe trm7Δ and trm734Δ mutants have more severe phenotypes than the S. cerevisiae mutants, and that tRNAPhe is the major biological target. Furthermore, we provide evidence that Trm7 and Trm732 function is widely conserved throughout eukaryotes, since human FTSJ1 and THADA, respectively, complement growth defects of S. cerevisiae trm7Δ and trm732Δ trm734Δ mutants by modifying C32 of tRNAPhe, each working with the corresponding S. cerevisiae partner protein. These results suggest widespread importance of 2′-O-methylation of the tRNA anticodon loop, implicate tRNAPhe as the crucial substrate, and suggest that this modification circuitry is important for human neuronal development.