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G-quadruplex-based CRISPR photoswitch for spatiotemporal control of genomic modulation
Huaping Deng1, Han Xu1, Yiru Wang1
1State Key Laboratory of Molecular Vaccinology and Molecular Diagnostics & Center for Molecular Imaging and Translational Medicine, School of Public Health, Xiamen University, Xiamen 361102, China.
Nucleic Acids Research
|March 13, 2023
Summary
Researchers developed a light-controlled CRISPR gene editing system using photoswitchable G-quadruplex guide RNA (GqRNA). This innovation enhances precision and reduces off-target effects for safer gene editing applications.
Area of Science:
- Molecular Biology
- Biotechnology
- Gene Editing Technologies
Background:
- CRISPR technology offers powerful gene regulation and editing capabilities.
- Precise control is crucial to mitigate off-target effects and excessive activity in CRISPR systems.
Purpose of the Study:
- To engineer a photoswitchable G-quadruplex guide RNA (GqRNA) for precise control of CRISPR-Cas9 gene editing.
- To enhance the stability, specificity, and safety of CRISPR gene editing through light-induced modulation.
Main Methods:
- Embedding dicationic azobenzene derivatives (AZD++) into G-quadruplex structures on crRNA (GqRNA).
- Utilizing light-induced isomerization of AZD++ to control GqRNA's active and inactive states.
- Evaluating GqRNA's stability, specificity, and editing efficiency in cellular contexts.
Main Results:
- G-quadruplex formation enhanced crRNA stability and sequence recognition specificity compared to sgRNA.
- Light activation of AZD++-GqRNA enabled rapid, efficient on-target genome editing.
- Photo-reversible switching of AZD++-GqRNA effectively inhibited cleavage and minimized off-target effects.
Conclusions:
- The developed photoswitchable GqRNA strategy offers precise, light-controlled modulation of CRISPR-Cas9 activity.
- This photo-reversible modality significantly improves the safety and applicability of CRISPR gene editing.
- The engineered system presents a novel approach for advanced gene therapy and research.
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