Synthetic mRNA Codon Optimization for Transcriptome-Wide Stability Control
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Solution Overview
Problem
Current methods fail to effectively modulate mRNA stability and protein expression across the entire transcriptome, as existing sequence and structural elements in untranslated regions cannot account for the wide variation in mRNA half-lives observed in Saccharomyces cerevisiae.
Innovation Solution
Modifying codon optimality in mRNA sequences by replacing optimal or non-optimal codons with synonymous codons to alter mRNA stability and protein expression levels, utilizing a set of specifically selected codons that influence mRNA degradation in a Dhh1p-dependent manner.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If existing sequence and structural elements in untranslated regions are used to regulate gene expression, then some control over protein expression is achieved, but the wide variation in mRNA half-lives observed in Saccharomyces cerevisiae cannot be effectively accounted for
Solution Approach 1:
The patent changes the parameter of codon optimality in mRNA sequences to modulate mRNA stability and protein expression. By replacing optimal or non-optimal codons with synonymous codons, the invention achieves transcriptome-wide modulation of mRNA half-lives, effectively accounting for the wide variation in stability observed in Saccharomyces cerevisiae that cannot be explained by untranslated region elements alone.
2Productivity
If codon optimality is modified to modulate mRNA stability, then significant modulation of protein expression levels is achieved, but the mRNA sequence must be synthetically modified
Solution Approach 1:
The patent creates synthetic copies of mRNA sequences with modified codon optimality. Instead of directly modifying existing mRNA in cells, the invention synthesizes new mRNA copies with predetermined codon compositions that encode the same protein sequence but exhibit different stability characteristics, enabling controlled modulation of protein expression levels.
Solution Approach 2:
The patent applies local quality changes by modifying specific codons within the mRNA coding sequence while maintaining the overall protein sequence. By selectively replacing individual optimal or non-optimal codons with synonymous alternatives, the invention locally alters mRNA stability properties without changing the encoded protein, achieving precise control over expression levels.
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 method achieves significant modulation of mRNA stability and protein expression levels, with synthetic mRNA sequences expressing proteins at least 10% different from wild-type sequences, demonstrating increased or decreased stability and expression based on codon modifications.
Implementation Method 1
Modifying codon optimality in mRNA sequences by replacing optimal or non-optimal codons with synonymous codons to alter mRNA stability and protein expression levels
Implementation Method 2
mRNA degradation plays a role in regulating transcript levels in the cell and is a major control point for modulating gene expression. Degradation of most mRNAs in Saccharomyces cerevisiae is initiated by removal of the 3′ poly(A) tail (deadenylation), followed by cleavage of the 5′ 7mGpppN cap (decapping) and exonucleolytic degradation of the mRNA body in a 5′-3′ direction
Data Source
AI summary
A synthetic cDNA which encodes a protein wherein at least one optimal or non-optimal codon in a wild type DNA encoding the protein has been replaced respectively with one or more non-optimal codons or optimal codons encoding the same amino acid.