Circular RNA Self-Splicing Design for Prolonged Protein Expression
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Solution Overview
Problem
The short half-life of messenger RNA (mRNA) in biological systems limits its effectiveness for therapeutic and engineering applications, necessitating a solution to extend protein expression duration.
Innovation Solution
A vector is designed to produce circular RNA (circRNA) within eukaryotic cells, comprising specific elements such as homology arms, group I intron fragments, and internal ribosome entry sites (IRES) to enhance stability and translation efficiency.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Duration of action of moving object
If messenger RNA is used for therapeutic applications, then protein expression can be achieved, but the half-life is relatively short limiting effectiveness
Solution Approach 1:
The patent applies the principle of curvature by converting linear mRNA into circular RNA structure. The circular configuration eliminates free ends that are susceptible to exonuclease degradation, thereby extending the half-life and stability of the RNA molecule in biological systems while maintaining its protein expression capability
Solution Approach 2:
The patent employs composite material principles by combining multiple functional elements within the circRNA structure, including IRES sequences for translation initiation, Kozak sequences for enhanced translation efficiency, and specific secondary structures that protect against degradation, creating a multifunctional RNA molecule with improved stability and expression duration
2Duration of action of stationary object
If circular RNA is produced using group I intron splicing, then RNA stability is extended, but the vector design becomes more complex
Solution Approach 1:
The patent applies segmentation by dividing the circRNA precursor into distinct functional modules: 5' homology arm, group I intron sequence, IRES element, coding sequence, and 3' homology arm. This modular design facilitates independent optimization of each element and simplifies the overall vector construction process
Solution Approach 2:
The patent uses preliminary action by incorporating group I intron sequences that automatically catalyze their own excision and circularization of the flanking RNA segments during transcription. This self-splicing mechanism occurs co-transcriptionally, eliminating the need for separate circularization steps and reducing experimental complexity
3Productivity
If IRES sequences are included to enhance translation, then translation efficiency improves, but the RNA structure becomes more complex
Solution Approach 1:
The patent applies universality by incorporating IRES sequences that provide translation initiation capability independent of the 5' cap structure. This allows the circRNA to utilize alternative translation pathways, enhancing translation efficiency while the modular design keeps the overall structure manageable through standardized functional elements
Applied Scientific Principles
This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.
Function Achieved in This Case
The vector allows for the production of biologically active circRNA that maintains prolonged protein expression and translation efficiency, enabling applications in therapeutic and diagnostic uses.
Implementation Method 1
a 3′ group I intron fragment containing a 3′ splice site dinucleotide... a 5′ group I intron fragment containing a 5′ splice site dinucleotide
Data Source
AI summary
Circular RNA and methods and constructs for engineering circular RNA are disclosed. In some embodiments, the circular RNA includes the following elements arranged in the following sequence: a) an adjacent exon sequence of a 3′ Group I self-splicing intron-exon, b) an internal ribosome entry site (IRES), c) a protein coding region or noncoding region, and d) an adjacent exon sequence of a 5′ Group I self-splicing intron-exon.