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Ribosome Profiling

Ribosome Profiling

Ribosome profiling or ribo-sequencing is a deep sequencing technique that produces a snapshot of active translation in a cell. It selectively sequences the mRNAs protected by ribosomes to get an insight into a cell’s translation landscape at any given point in time.
Applications of ribosome profiling
Ribosome profiling has many applications, including in vivo monitoring of translation inside a particular organ or tissue type and quantifying new protein synthesis levels.
The technique helps...

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Simultaneous Affinity Enrichment of Two Post-Translational Modifications for Quantification and Site Localization

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Spatial Mapping and Interactome Profiling of m⁶A-Modified R-Loops via Chemically Inducible Split-APEX2 Proximity

Meng-Die Zhu¹, Hua-Qian Yin¹, Zhong-Da He¹

¹State Key Laboratory of Medicinal Chemical Biology, Frontiers Science Centre for New Organic Matter, Tianjin Key Laboratory of Biosensing and Molecular Recognition, Research Centre for Analytical Sciences, College of Chemistry, School of Medicine and Frontiers Science Centre for Cell Responses, Nankai University, Tianjin 300071, P. R. China.

Analytical Chemistry

|December 3, 2025

Summary

Researchers developed m6A-R-loop proximity labeling (m6A-RLPL) to visualize RNA modifications in live cells. This method revealed m6A-R-loops in nucleoli and viral replication compartments, identifying key protein regulators.

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Area of Science:

Molecular Biology
Epigenetics
Virology

Background:

N6-methyladenosine (m6A)-modified R-loops are critical for epigenetic regulation and genome stability.
Understanding their spatial distribution and protein interactions in living cells is challenging.

Purpose of the Study:

To develop a novel proximity labeling technology for spatially resolved profiling of m6A-R-loops and their interactomes in live cells.
To investigate the dynamics and organization of m6A-R-loops during host and viral processes.

Main Methods:

Development of m6A-R-loop proximity labeling (m6A-RLPL) using a chemically inducible split-APEX2 system.
Integration of dual-target recognition (R-loop and m6A) with ABA-inducible dimerization for signal amplification.
Application in live cells to study host m6A-R-loops and herpes simplex virus (HSV) infection.

Main Results:

m6A-RLPL revealed host m6A-R-loops are enriched in nucleoli under normal conditions.
Viral m6A-R-loops were observed accumulating in phase-separated granules within HSV replication compartments during late-stage infection.
Proximity proteomics identified ZC3H4 and CCDC124 as regulators of these viral m6A-R-loop structures, which function as transcription sites for HSV late genes.

Conclusions:

m6A-RLPL provides a powerful, generalizable tool for studying m6A-R-loop organization and interactions in living systems.
The study elucidates the role of m6A-R-loops in viral gene expression and identifies novel regulatory factors.