Life without post-transcriptional addition of G(−1): two alternatives for tRNA(His) identity in Eukarya

The identity of tRNA(His) is strongly associated with the presence of an additional 5′-guanosine residue (G(−1)) in all three domains of life. The critical nature of the G(−1) residue is underscored by the fact that two entirely distinct mechanisms for its acquisition are observed, with cotranscriptional incorporation observed in Bacteria, while post-transcriptional addition of G(−1) occurs in Eukarya. Here, through our investigation of eukaryotes that lack obvious homologs of the post-transcriptional G(−1)-addition enzyme Thg1, we identify alternative pathways to tRNA(His) identity that controvert these well-established rules. We demonstrate that Trypanosoma brucei, like Acanthamoeba castellanii, lacks the G(−1) identity element on tRNA(His) and utilizes a noncanonical G(−1)-independent histidyl-tRNA synthetase (HisRS). Purified HisRS enzymes from A. castellanii and T. brucei exhibit a mechanism of tRNA(His) recognition that is distinct from canonical G(−1)-dependent synthetases. Moreover, noncanonical HisRS enzymes genetically complement the loss of THG1 in Saccharomyces cerevisiae, demonstrating the biological relevance of the G(−1)-independent aminoacylation activity. In contrast, in Caenorhabditis elegans, which is another Thg1-independent eukaryote, the G(−1) residue is maintained, but here its acquisition is noncanonical. In this case, the G(−1) is encoded and apparently retained after 5′ end processing, which has so far only been observed in Bacteria and organelles. Collectively, these observations unearth a widespread and previously unappreciated diversity in eukaryotic tRNA(His) identity mechanisms.