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Single-Molecule Fluorescence Visualization of DNA Polymerase Dynamics at G-Quadruplexes
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Single-molecule fluorescence studies on cotranscriptional G-quadruplex formation coupled with R-loop formation
Gunhyoung Lim1, Sungchul Hohng1
1Department of Physics and Astronomy, and Institute of Applied Physics, Seoul National University, Seoul 08826, Republic of Korea.
Nucleic Acids Research
|August 19, 2020
Summary
G-quadruplex (GQ) formation during transcription is rare but facilitated by R-loops. Stable GQs accumulate and promote further R-loop formation, creating a positive feedback loop impacting gene regulation and genome stability.
Area of Science:
- Molecular Biology
- Genetics
- Biophysics
Background:
- G-quadruplexes (GQ) are DNA secondary structures found in gene regulatory regions and telomeres.
- GQ structures are implicated in gene regulation and genome instability.
- Mechanisms of GQ formation, particularly during transcription, remain unclear.
Purpose of the Study:
- To investigate the cotranscriptional formation of G-quadruplexes (GQ) coupled with R-loop formation.
- To elucidate the interplay and feedback mechanisms between GQ and R-loop formation during transcription.
Main Methods:
- Single-molecule fluorescence experiments were employed.
- T7 RNA polymerase was used to monitor cotranscriptional GQ and R-loop formation.
- RNase H treatment was used to assess GQ stability.
Main Results:
- GQ formation occurred rarely per transcription round but was preceded and facilitated by R-loop formation.
- Some GQs exhibited extreme stability, persisting through RNase H treatment and accumulating over multiple transcription rounds.
- GQ on the non-template strand enhanced R-loop formation in subsequent transcription rounds.
Conclusions:
- A positive feedback mechanism exists between G-quadruplex and R-loop formation during transcription.
- This feedback loop may play a significant role in gene regulation and genome instability.
- Understanding these mechanisms offers insights into potential therapeutic targets.
