Nature-Inspired Chemical Probes for Labeling tRNAs Containing 2-Thiouridine (s²U)

Area of Science:

• Biochemistry
• Molecular Biology
• Chemical Biology

Background:

• RNA modifications are crucial for cellular functions and disease.
• Hydrophobic prenyl-modifications exist in RNA, distinct from common methylations.
• Specific RNA modifications, like 2-thiouridine (s²U), are implicated in cellular processes.

Purpose of the Study:

• To develop a novel chemical labeling strategy for specific RNA tagging.
• To investigate the biological roles of 2-thiouridine (s²U) containing tRNAs.
• To enable visualization and monitoring of s²U-tRNAs in cellular environments.

Main Methods:

• Synthesis of "clickable" geranyl-pyrophosphate analogs.
• Utilizing a natural SelU-mediated geranylation process for tRNA tagging.
• A two-step azidation-and-click-tagging (ACT-Flu) approach with fluorescent dyes.

Main Results:

• Identification of two effective geranyl-pyrophosphate analogs for labeling.
• Successful application of the ACT-Flu method for in-cell RNA imaging.
• Demonstration of ACT-Flu's utility in studying s²U-tRNAs in healthy and diseased cells.

Conclusions:

• The ACT-Flu method provides a powerful toolset for biochemical studies of RNA modifications.
• This technique facilitates the monitoring of s²U-tRNA status in various cellular conditions.
• The findings advance our understanding of RNA modifications' roles in human health and disease.