Eukaryotic life without tQCUG: the role of Elongator-dependent tRNA modifications in Dictyostelium discoideum

Abstract

<jats:title>Abstract</jats:title> <jats:p>In the Elongator-dependent modification pathway, chemical modifications are introduced at the wobble uridines at position 34 in transfer RNAs (tRNAs), which serve to optimize codon translation rates. Here, we show that this three-step modification pathway exists in Dictyostelium discoideum, model of the evolutionary superfamily Amoebozoa. Not only are previously established modifications observable by mass spectrometry in strains with the most conserved genes of each step deleted, but also additional modifications are detected, indicating a certain plasticity of the pathway in the amoeba. Unlike described for yeast, D. discoideum allows for an unconditional deletion of the single tQCUG gene, as long as the Elongator-dependent modification pathway is intact. In gene deletion strains of the modification pathway, protein amounts are significantly reduced as shown by flow cytometry and Western blotting, using strains expressing different glutamine leader constructs fused to GFP. Most dramatic are these effects, when the tQCUG gene is deleted, or Elp3, the catalytic component of the Elongator complex is missing. In addition, Elp3 is the most strongly conserved protein of the modification pathway, as our phylogenetic analysis reveals. The implications of this observation are discussed with respect to the evolutionary age of the components acting in the Elongator-dependent modification pathway.</jats:p>

Experience with German Research Consortia in the Field of Chemical Biology of Native Nucleic Acid Modifications

“…Functional analyses of tRNA modifications focused conspicuously on the so-called wobble modifications at position 34, ,− ,− as well as on changes in tRNA modifications in response to stress, especially with reference to nutrient supply (“nutritional stress” , ). Interestingly, some modifications that vary under such stress have been shown to influence recognition by the human innate immune system, a function that may be directly relevant to the development of RNA-based vaccines.…”

Changes of the tRNA Modification Pattern during the Development of Dictyostelium discoideum

“…Yet, analysis of the RT signatures alone cannot exclude the possibility of misclassifications or false positive hits resulting from, e.g., robust secondary structures. To be able to classify the modifications unambiguously, specific chemical treated RNA-seq data or mass spectrometry analyses would be necessary [35,52,53]. Here, we are content with well-founded hypotheses concerning the identity of the modifications and focus on the variations of modification patterns across the developmental time course.…”

Recent advances in the molecular understanding of medulloblastoma

“…Therefore, loss‐of‐function of ELP1 impairs elongator‐dependent tRNA modification at the wobble position. The chemical modification of uridines at the wobble position is critical for proper mRNA decoding, and its absence influences codon translation rates 44,45 . Therefore, mutant ELP1 tumors have a significant codon usage bias, where AA‐ending codons are inefficiently recognized but AG‐ending codons are efficiently recognized (Figure 3B).…”

“…The chemical modification of uridines at the wobble position is critical for proper mRNA decoding, and its absence influences codon translation rates. 44 , 45 Therefore, mutant ELP1 tumors have a significant codon usage bias, where AA‐ending codons are inefficiently recognized but AG‐ending codons are efficiently recognized (Figure 3B ). 7 , 46 The shift in codon usage gives rise to the significant upregulation of gene sets associated with RNA splicing, amino acid activation, and activation of the endoplasmic reticulum stress pathway in SHH medulloblastoma with ELP1 mutation.…”