Where have all the inosines gone? Conflicting evidence for A‐to‐I editing of the anticodon of higher eukaryotic questions the dogma of a universal wobble‐mediated decoding of CGN codons

Abstract

Codon-anticodon recognition between triplets of an mRNA and a specific tRNA is the key element in the translation of the genetic code. In general, the precision of this process is dominated by a strict Watson-Crick base-pairing scheme. However, the degeneracy of the genetic code led Crick to propose the Wobble Hypothesis, permitting a less restraining interaction with the third base of the codon and involving the participation of inosine for decoding C-ending codons. The concept that the anticodon base A34 of tRNA Arg ACG in all eukaryotes, eubacteria, and plant chloroplasts is converted to I34 is firmly anchored in the literature despite conflicting evidence for its existence in higher eukaryote cytoplasmic tRNA Arg ACG . Here, we provide additional data and summarize the arguments favoring and contradicting post-transcriptional deamination of this position. A hypothesis that resolves the apparent conflict is proposed.

Detection of Inosine on Transfer RNAs without a Reverse Transcription Reaction

“…For example, while ∼90% of sequencing reads mapping to tRNA Val AAC reflected the presence of I34, <2% of reads mapping to tRNA Arg ACG evidenced I34 modification on this tRNA . This is consistent with other reports that describe the difficulty of detecting I34 on tRNA Arg ACG in higher eukaryotes by RT-based methods …”

“…I34 is an essential posttranscriptional tRNA modification of biomedical importance that played an important role in the evolution of eukaryotic genomes and proteomes , Currently available methods for detecting I34, such as RT-based sequencing strategies, can be limited by technical artifacts that compromise their interpretation (Figure ). We have previously shown that deep sequencing of tRNAs can be used to detect I34 on precursor tRNAs in vivo , but the efficiency of I34 detection varied considerably among different I34-tRNA species.…”

“…In recent years, conflicting evidence has emerged regarding the I34 modification status of tRNA Arg ACG in higher eukaryotes . While I34 could be readily detected by sequencing of tRNA Arg ACG in Escherichia coli , Mycoplasma capricolum , and Saccharomyces cerevisiae , it could not be detected in nematodes, mosses, higher plants, insects, or mammals. − However, the presence of I34 on human tRNA Arg ACG has been documented by other means, suggesting that at least in some of these cases RT-based methods may be leading to artifactual results.…”

“…Given the difficulty in detecting I34 on human tRNA Arg ACG by RT-based methods (Figure ), , we next applied SL-ID to verify the presence of I34 on this tRNA in HEK293T cells. We found that A34 is modified to I34 on this tRNA and that the modified base is efficiently and specifically identified by SL-ID (Figure C).…”

Chimeric mitochondrial peptides from contiguous regular and swinger RNA

“…These analyses show that detected peptides are more likely translated according to the mitochondrial genetic code than according to the nuclear genetic code. Note that translation, within the mitochondrion, according to the nuclear code is possible: it potentially depends for some codons upon the presence of cytosolic tRNAs, which could be occasionally imported in mitochondria [21] , [32] , [39] , [44] , [75] , [76] , [78] , [112] , [114] . However, this rationale is not symmetric: cytosolic translation according to the mitochondrial genetic code is much less probable than the opposite, so that nuclear origins are not compatible with the results obtained.…”