YAP1 and TAZ RNA Base Editing With Guide RNA Precision
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Solution Overview
Problem
Current RNA base editing technologies lack specificity and efficiency, leading to off-target effects and challenges in precisely targeting ADAR enzymes to modify coding potential of mRNAs, particularly for therapeutic applications in genetic and non-genetic diseases.
Innovation Solution
Employing guide RNAs to direct catalytically inactive Cas proteins, such as dCas13, to specific sites on mRNA encoding YAP1 or TAZ, enabling precise A-to-I or C-to-U editing, thereby altering post-translational modification sites to regulate kinase signaling pathways and promote therapeutic benefits.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If ADAR enzymes are used for RNA base editing, then editing activity is achieved, but specificity and targeting precision are insufficient leading to off-target effects
Solution Approach 1:
The patent introduces guide RNAs as intermediary molecules that mediate between the ADAR enzyme and the target RNA. The guide RNA contains a guide sequence that is complementary to the target site, enabling precise positioning of the ADAR enzyme at the desired location on the RNA molecule, thereby achieving specific base editing while minimizing off-target effects
Solution Approach 2:
The patent applies local quality by designing guide RNAs with specific guide sequences that are complementary to particular target sites on the RNA. This allows the editing activity to be concentrated at specific locations rather than occurring throughout the entire RNA molecule, achieving localized base editing with high precision
2Reliability
If DNA editing strategies are used, then permanent genetic modification is achieved, but risk of permanent off-target mutations in the genome increases
Solution Approach 1:
The patent extracts the editing process from the DNA level and applies it at the RNA level. By using ADAR enzymes to edit RNA molecules directly, the invention achieves permanent functional modification of the protein product without making permanent changes to the genomic DNA, thereby eliminating the risk of permanent off-target mutations in the genome
Solution Approach 2:
The patent uses RNA as a temporary copy of the genetic information. By editing the RNA copy rather than the original DNA template, the invention achieves functional modification without permanently altering the genome. The edited RNA is then translated into modified protein, achieving the desired effect without permanent DNA changes
3Duration of action of moving object
If RNA editing is used, then reversible and controlled editing is achieved, but efficiency and specificity of base editing remain insufficient
Solution Approach 1:
The patent employs guide RNAs as intermediaries that enhance the efficiency of RNA editing by directing ADAR enzymes to specific target sites. The guide RNA-mediated approach increases editing efficiency while maintaining reversibility, as the edited RNA molecules can be degraded and new unedited RNA can be transcribed from the original DNA template
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
This approach allows for reversible and controlled RNA editing, minimizing off-target effects and providing a novel regenerative therapy for cardiac disease by activating YAP1 and TAZ proteins, enhancing cell proliferation and tissue regeneration.
Implementation Method 1
guide RNAs for precisely directing deaminase enzymes to specific sites on an mRNA
Implementation Method 2
recruitment of adenosine deaminases acting on RNA (ADAR) enzymes to RNA. ADARs recognize adenosine on RNA
Data Source
AI summary
The present invention provides novel RNA base editing compositions, systems, methods and uses. Guide RNAs for site-specific RNA editing of RNA encoding transcriptional coactivators YAP1 or TAZ are provided, and compositions and systems comprising the same with a programmable RNA binding protein (e.g. a Cas protein) and/or a base editor. Methods for RNA editing of YAP1 or TAZ are also provided. RNA editing of YAP1 or TAZ is used for targeting phosphorylation sites, and activating transcription of proteins in regenerative therapy for treating cardiac disease.