Conserved 5-methyluridine tRNA modification modulates ribosome translocation.

While the centrality of posttranscriptional modifications to RNA biology has long been acknowledged, the function of the vast majority of modified sites remains to be discovered. Illustrative of this, there is not yet a discrete biological role assigned for one of the most highly conserved modifications, 5-methyluridine at position 54 in tRNAs (mU54). Here, we uncover contributions of mU54 to both tRNA maturation and protein synthesis. Our mass spectrometry analyses demonstrate that cells lacking the enzyme that installs mU in the T-loop (TrmA in , Trm2 in ) exhibit altered tRNA modification patterns. Furthermore, mU54-deficient tRNAs are desensitized to small molecules that prevent translocation in vitro. This finding is consistent with our observations that relative to wild-type cells, cell growth and transcriptome-wide gene expression are less perturbed by translocation inhibitors. Together our data suggest a model in which mU54 acts as an important modulator of tRNA maturation and translocation of the ribosome during protein synthesis.