Global translational impacts of the loss of the tRNA modification tA in yeast.

The universal tRNA modification tA is found at position 37 of nearly all tRNAs decoding ANN codons. The absence of tA leads to severe growth defects in baker's yeast, phenotypes similar to those caused by defects in mcmsU synthesis. Mutants in mcmsU can be suppressed by overexpression of tRNA, but we show tA phenotypes could not be suppressed by expressing any individual ANN decoding tRNA, and tA and mcmsU are not determinants for each other's formation. Our results suggest that tA deficiency, like mcmsU deficiency, leads to protein folding defects, and show that the absence of tA led to stress sensitivities (heat, ethanol, salt) and sensitivity to TOR pathway inhibitors. Additionally, L-homoserine suppressed the slow growth phenotype seen in tA-deficient strains, and proteins aggregates and Advanced Glycation End-products (AGEs) were increased in the mutants. The global consequences on translation caused by tA absence were examined by ribosome profiling. Interestingly, the absence of tA did not lead to global translation defects, but did increase translation initiation at upstream non-AUG codons and increased frame-shifting in specific genes. Analysis of codon occupancy rates suggests that one of the major roles of tA is to homogenize the process of elongation by slowing the elongation rate at codons decoded by high abundance tRNAs and I:C pairs while increasing the elongation rate of rare tRNAs and G:U pairs. This work reveals that the consequences of tA absence are complex and multilayered and has set the stage to elucidate the molecular basis of the observed phenotypes.