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Defining expansions and perturbations to the RNA polymerase III transcriptome and epitranscriptome by modified direct
Ruth Verstraten1,2, Pierina Cetraro1, Amy H Fitzpatrick1,3
1Institute of Virology, Hannover Medical School, Hannover, Germany.
Nature Communications
|January 6, 2026
Summary
A new method, DRAP3R, analyzes RNA polymerase III (Pol III) transcribed RNAs and their modifications. This tool expands the understanding of the Pol III transcriptome and epitranscriptome, revealing novel RNAs and modification patterns.
Area of Science:
- Molecular Biology
- Genomics
- RNA Biology
Background:
- RNA polymerase III (Pol III) transcribes essential non-coding RNAs, but analysis methods are limited.
- Understanding Pol III transcription, processing, and RNA modifications is crucial for cellular function.
Purpose of the Study:
- To develop a sensitive and systematic method for analyzing Pol III transcribed RNAs and their modifications.
- To expand the known Pol III transcriptome and characterize RNA modifications at single-nucleotide resolution.
Main Methods:
- Developed DRAP3R (Direct Read and Analysis of Polymerase III transcribed RNAs), a modified nanopore direct RNA sequencing approach.
- Applied DRAP3R to different cell types, including those infected with Herpes Simplex Virus Type 1.
Main Results:
- Identified previously unconfirmed tRNA genes and novel Pol III transcribed RNAs, expanding the Pol III transcriptome.
- Enabled discrimination of RNA modifications like pseudouridine (Ψ) and N6-methyladenosine (m6A) at single-nucleotide resolution.
- Revealed differential Ψ installation patterns and extensive Pol III transcriptome/epitranscriptome remodeling during viral infection.
Conclusions:
- DRAP3R is a powerful tool for systematic study of Pol III transcribed RNAs and their modifications.
- The study expands the understanding of the Pol III transcriptome and epitranscriptome in various cellular contexts.
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