Reversible oxidative dimerization of 4-thiouridines in tRNA isolates

Area of Science:

• Biochemistry
• Molecular Biology
• RNA Research

Background:

• Non-canonical nucleoside structures in RNA are gaining attention for their roles in RNA modifications and RNA vaccines.
• These modifications add functionality to RNA biopolymers, which are otherwise chemically homogenous.
• The discovery of novel RNA structures is crucial for understanding RNA function and applications.

Purpose of the Study:

• To identify and characterize a presumed novel RNA modification.
• To investigate the spontaneous formation of disulfide-linked 4-thiouridine (s⁴U) dimers in E. coli tRNA.
• To understand the implications of artifactual modifications in RNA research.

Main Methods:

• Liquid chromatography-tandem mass spectrometry (LC-MS/MS) was employed for identification.
• Stable isotope labeling was used to trace the origin of the modification.
• Analysis focused on isolates of *E. coli* tRNA.

Main Results:

• A disulfide-linked dimer of 4-thiouridine (s⁴U) was identified as a spontaneous modification in *E. coli* tRNA.
• Stable isotope labeling indicated the dimer formed *ex vivo* via oxidation by ambient oxygen.
• The findings suggest this is not a novel *in vivo* modification but an artifact of sample handling.

Conclusions:

• The study underscores the importance of caution in identifying new RNA modifications due to potential artifacts.
• It raises awareness of RNA vulnerability to oxidative damage during transport and storage.
• Proper handling and storage protocols are essential to prevent artifactual RNA modifications.