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Upgraded circular single-stranded DNA regulators for multiple-input multiple-output gene circuits in mammalian cells
Linlin Tang1, Jinghao Wang1,2, Kaiqi Xu1,2
1Hangzhou Institute of Medicine, Chinese Academy of Sciences, Zhejiang, Hangzhou, 310022, China.
Science Advances
|May 16, 2025
Summary
Researchers developed a new gene-editing tool using engineered circular single-stranded DNA (cssDNA) regulators with "bridge" designs. This innovation enhances control over gene expression for advanced synthetic gene networks in genetic diagnostics and therapy.
Area of Science:
- Synthetic Biology
- Molecular Biology
- Genetic Engineering
Background:
- Synthetic gene networks are crucial for diagnostics and therapy but limited by molecular tools.
- Previous circular single-stranded DNA (cssDNA) regulators had restricted flexibility and lacked multi-input capabilities.
- Existing systems struggled with endogenous orthogonal regulation.
Purpose of the Study:
- To engineer a novel "bridge" design for cssDNA regulators.
- To overcome limitations of previous cssDNA systems, enabling greater flexibility and multi-input control.
- To create a versatile platform for advanced synthetic gene networks.
Main Methods:
- Engineered "bridge" designs into cssDNA regulators.
- Developed sequence-programmable switches responsive to endogenous signals (ATP, APE1, RNase H).
- Utilized cssDNA orthogonality to construct three-input, three-output genetic circuits.
Main Results:
- The new bridge design enhanced regulatory sequence flexibility.
- Achieved multiple-input, multiple-output (MIMO) signal control.
- Enabled trans-regulation within and between cssDNAs, demonstrating endogenous orthogonal regulation.
- Successfully constructed complex three-input, three-output genetic circuits.
Conclusions:
- The upgraded cssDNA regulatory strategy offers a versatile and powerful platform for gene regulation.
- This approach overcomes key limitations of previous systems, paving the way for sophisticated synthetic gene networks.
- The engineered cssDNA regulators show promise for advancing genetic diagnostics and gene therapy applications.
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