Split RNA switch orchestrates pre- and post-translational control to enable cell type-specific gene expression

Area of Science:

• Synthetic biology
• Molecular and cell biology
• Biotechnology

Background:

• RNA switches offer precise gene expression control but often lack specificity.
• Low ON/OFF ratios and target identification challenges limit practical applications of individual RNA switches.

Purpose of the Study:

• To develop "split RNA switches" that integrate multiple RNA switches via protein splicing to enhance specificity and ON/OFF ratios.
• To demonstrate the utility of split RNA switches in cell purification, genome editing, and synthetic RNA-based circuits.

Main Methods:

• Exploitation of protein splicing to create split RNA switches.
• Development of microRNA-responsive ON switch systems with improved ON/OFF ratios.
• Application in cell purification using drug-resistance genes and CRISPR-mediated genome editing.
• Construction of RNA-based synthetic circuits for detecting multiple microRNAs and proteins with logical operations.

Main Results:

• Split RNA switches significantly improved the ON/OFF ratio of microRNA-responsive systems by reducing leaky expression in human cells.
• Efficient cell purification and CRISPR-mediated genome editing with minimal off-target effects were achieved using endogenous microRNA profiles.
• Demonstrated RNA-based synthetic circuits capable of detecting multiple microRNAs and proteins through logical operations.

Conclusions:

• Split RNA switches represent a significant advancement in mRNA-based therapeutic technologies by leveraging post-translational processing.
• This technology enhances specificity and ON/OFF ratios, overcoming limitations of individual RNA switches.
• Split RNA switches hold promise for diverse applications including gene therapy, regenerative medicine, and advanced synthetic biology circuits.