Dual pathways of tRNA hydroxylation ensure efficient translation by expanding decoding capability

Area of Science:

• Molecular Biology
• Biochemistry
• Genetics

Background:

• Bacterial tRNAs feature modified nucleosides like 5-carboxymethoxyuridine (cmo5U) at the anticodon's first position.
• These modifications enable non-Watson-Crick base pairing, expanding codon decoding capabilities.
• The biogenesis and physiological significance of cmo5U modifications were previously unclear.

Purpose of the Study:

• To investigate the biogenesis of 5-hydroxyuridine (ho5U), a precursor to cmo5U.
• To identify the enzymes responsible for ho5U formation.
• To elucidate the physiological roles of tRNA hydroxylation in bacterial decoding.

Main Methods:

• Employed reverse genetics approaches to study gene functions.
• Utilized comparative genomics to identify conserved genetic elements.
• Generated and analyzed E. coli strains deficient in specific genes (trhP and trhO).

Main Results:

• Identified TrhP (YegQ) and TrhO (YceA) as essential enzymes for ho5U formation.
• TrhP mediates prephenate-dependent ho5U synthesis, while TrhO catalyzes oxygen-dependent synthesis.
• E. coli strains lacking both trhP and trhO showed temperature sensitivity and impaired decoding of G-ending codons.

Conclusions:

• TrhP and TrhO are critical for the biogenesis of modified uridine nucleosides in bacterial tRNA.
• tRNA hydroxylation is essential for efficient and accurate decoding of specific codons during protein synthesis.
• The identified pathways highlight the importance of post-transcriptional tRNA modifications for bacterial physiology.