The Establishment of Artificial RNA Cascade Circuits for Gene Regulation Based on Doxycycline-Induced Pre-mRNA Alternative Splicing
Guimin Dai, Jiawen Cheng, Weiran Liu, Xueli Yin, Yuanyuan Zhang
TL;DR
This paper describes a new method using RNA switches to control gene expression in mammalian cells with doxycycline, enabling precise and reversible regulation.
Contribution
The novel contribution is the development of RNA cascade circuits using doxycycline-induced pre-mRNA splicing for dynamic gene regulation.
Findings
A chimeric intron module with RNA riboswitch and TetR aptamer was developed to regulate gene expression via doxycycline.
Two contrasting switches (CTM and C2ITetR>4A) were created to reversibly control gene expression and tumor cell apoptosis.
An RNA cascade system was designed to minimize crosstalk and enhance gene expression control using a single ligand.
Abstract
This study developed an artificial chimeric intron module with an RNA riboswitch and TetR aptamer that were integrated into essential gene exons. Doxycycline can modulate Pre-mRNA alternative splicing, modify the exon reading frame, and dynamically regulate gene expression. By shifting the aptamer 2 base pair within the switch, we unexpectedly obtained the “on-switch” CTM and “off-switch” C2ITetR>4A, which possess thoroughly contrasting regulatory functions. The CTM module can conditionally induce tumor cell apoptosis and regulate genes reversibly and sustainably following doxycycline induction. We integrated the C2ITetR>4A/CTM switches with the L7Ae/k-turn module to create an intron-spliced double-switched RNA cascade system. The system can both activate and inhibit the splicing mechanism utilizing the same ligand to minimize crosstalk among aptamer switching elements, control target…
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Taxonomy
TopicsRNA and protein synthesis mechanisms · Advanced biosensing and bioanalysis techniques · RNA Research and Splicing