ArticleInternational journal of molecular sciences2025
The Establishment of Artificial RNA Cascade Circuits for Gene Regulation Based on Doxycycline-Induced Pre-mRNA Alternative Splicing.
Article in International journal of molecular sciences, 2025. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Not yet cited in PubMed.
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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 gene leakage, and enhance the dynamic range of gene expression. We analyzed numerous factors affecting Pre-mRNA splicing to identify the optimal equilibrium point for switch regulation. This will enable precise predictions of dynamic regulatory efficiency and the rational design of genetic modules, thereby providing a valuable instrument for mammalian synthetic biology.
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