Programmable RNA Methylation Editors for Precise Epigenetic Targeting
Find Innovative SolutionsGenerate Solutions
Solution Overview
Problem
Current methods lack the ability to efficiently and specifically target the addition or removal of methylation sites in RNA, which are crucial for understanding and manipulating epigenetic changes associated with cellular processes and diseases.
Innovation Solution
Development of RNA programmable methylation 'writers' and demethylation 'erasers' in the form of fusion proteins, which can install or remove methyl groups in RNA molecules with high specificity and efficiency, operating in both the nucleus and cytoplasm.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If conventional methods are used to detect or modify RNA methylation, then the process is non-specific and inefficient, but the patent develops RNA programmable fusion proteins that achieve high specificity and efficiency in targeting methylation sites
Solution Approach 1:
The patent uses guide RNA as an intermediary molecule that bridges the Cas13b protein and the target RNA sequence. The guide RNA specifically binds to the target sequence through base pairing, directing the fusion protein to the precise methylation site without requiring complex protein-protein recognition systems.
Solution Approach 2:
The patent merges multiple functional domains into a single fusion protein: the Cas13b RNA-binding domain for target recognition, the methyltransferase domain for methylation, and the demethylase domain for demethylation. This consolidation achieves high specificity through the guide RNA while maintaining operational simplicity.
2Manufacturing precision
If RNA methylation editing is performed without programmable targeting, then the process is simple, but it lacks the ability to specifically target particular methylation sites in RNA molecules
Solution Approach 1:
The patent employs a dynamic and adaptable guide RNA sequence that can be easily reconfigured to target different RNA sequences. By simply changing the guide RNA sequence rather than redesigning the entire protein system, the method achieves high precision targeting while maintaining ease of operation.
Solution Approach 2:
The system separates the targeting function (guide RNA) from the catalytic function (fusion protein). This segmentation allows independent optimization: the guide RNA provides programmable specificity while the fusion protein provides robust catalytic activity, making the overall system both precise and easy to operate.
3Stability of the object's composition
If traditional epigenetic modification methods are used, then the nucleotide sequence must be changed, but the patent achieves epigenetic modification without altering the underlying nucleotide sequence
Solution Approach 1:
The patent converts the transient nature of RNA into a benefit: by modifying RNA methylation states rather than DNA sequences, the system achieves reversible and controllable epigenetic effects. The RNA modifications can be dynamically adjusted without permanent genetic changes, providing reliable and tunable epigenetic control.
Solution Approach 2:
The patent changes the methylation state parameter of RNA molecules (adding or removing methyl groups) without altering the fundamental nucleotide sequence. This parameter change approach allows epigenetic modification while preserving the stability and integrity of the underlying genetic code.
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
These fusion proteins enable precise editing of RNA methylation states, affecting RNA stability, expression, and splicing, offering potential therapeutic applications for diseases like cancer by altering epigenetic conditions without changing the nucleotide sequence.
Implementation Method 1
an RNA-programmable RNA binding domain
Implementation Method 2
an effector domain, wherein the effector domain is capable of adding or removing a methyl group in an RNA
Implementation Method 3
effector domain is capable of adding or removing a methyl group
Data Source
AI summary
The disclosure provides programmable methylation “writers” and demethylation “erasers” for editing the methylation state of RNA targets, e.g., an RNA transcriptome. In particular, the disclosure provides RNA methylation editor polynucleotide constructs and vectors comprising (i) an RNA programmable RNA binding domain (RNApRNAbd); and (ii) an effector domain, wherein the effector domain is capable of adding or removing a methyl group in an RNA. The disclosed RNA methylation editor constructs are capable of achieving limited off-target modifications in RNA molecules. Further, the disclosure provides methods for making and using the programmable methylation editors to modifying the methylation state of RNA. The disclosure further provides complexes comprising a methylation writer protein and a guide RNA molecule and complexes comprising a demethylation eraser protein and a guide RNA molecule. The disclosure further provides pharmaceutical compositions and cells comprising the disclosed fusion proteins and complexes.