Eukaryotic AlaX provides multiple checkpoints for quality and quantity of aminoacyl-tRNAs in translation.
Nucleic Acids Res
; 52(13): 7825-7842, 2024 Jul 22.
Article
in En
| MEDLINE
| ID: mdl-38869066
ABSTRACT
Translational fidelity relies critically on correct aminoacyl-tRNA supply. The trans-editing factor AlaX predominantly hydrolyzes Ser-tRNAAla, functioning as a third sieve of alanyl-tRNA synthetase (AlaRS). Despite extensive studies in bacteria and archaea, the mechanism of trans-editing in mammals remains largely unknown. Here, we show that human AlaX (hAlaX), which is exclusively distributed in the cytoplasm, is an active trans-editing factor with strict Ser-specificity. In vitro, both hAlaX and yeast AlaX (ScAlaX) were capable of hydrolyzing nearly all Ser-mischarged cytoplasmic and mitochondrial tRNAs; and robustly edited cognate Ser-charged cytoplasmic and mitochondrial tRNASers. In vivo or cell-based studies revealed that loss of ScAlaX or hAlaX readily induced Ala- and Thr-to-Ser misincorporation. Overexpression of hAlaX impeded the decoding efficiency of consecutive Ser codons, implying its regulatory role in Ser codon decoding. Remarkably, yeast cells with ScAlaX deletion responded differently to translation inhibitor treatment, with a gain in geneticin resistance, but sensitivity to cycloheximide, both of which were rescued by editing-capable ScAlaX, alanyl- or threonyl-tRNA synthetase. Altogether, our results demonstrated the previously undescribed editing peculiarities of eukaryotic AlaXs, which provide multiple checkpoints to maintain the speed and fidelity of genetic decoding.
Full text:
1
Collection:
01-internacional
Database:
MEDLINE
Main subject:
Saccharomyces cerevisiae
/
Protein Biosynthesis
/
Alanine-tRNA Ligase
Limits:
Humans
Language:
En
Journal:
Nucleic Acids Res
Year:
2024
Document type:
Article
Affiliation country:
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