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Engineering Small Molecule Switches of Protein Function in Zebrafish Embryos
Wes Brown1, Joshua Wesalo1, Michael Tsang2
1Department of Chemistry, University of Pittsburgh, Pittsburgh, Pennsylvania 15260, United States.
Journal of the American Chemical Society
|January 20, 2023
Summary
Scientists developed a new method using genetic code expansion to control protein activity in zebrafish embryos with precise timing. This technique allows for targeted chemical intervention during development, offering new insights into congenital heart defects and enzymatic processes.
Area of Science:
- Developmental Biology
- Chemical Biology
- Molecular Biology
Background:
- Precise temporal regulation of protein function is crucial for complex developmental processes.
- Zebrafish embryos are a valuable model organism for studying development.
- Limited availability of general small molecule switches for protein function in zebrafish hinders targeted interventions.
Purpose of the Study:
- To develop and apply a broadly applicable method for temporally controlled protein activation in zebrafish embryos.
- To investigate the role of specific proteins in developmental events using conditional activation.
- To explore the potential of small molecule-triggered protein function for studying congenital heart defects and enzymatic activities.
Main Methods:
- Utilized genetic code expansion for site-specific incorporation of unnatural amino acids with caging groups.
- Employed small molecule triggers (phosphines or tetrazines) to remove caging groups and activate proteins.
- Applied this method to activate enzymes like GTPases and proteases in zebrafish embryos.
- Investigated a RASopathy mutant of NRAS to study congenital heart defect development.
Main Results:
- Achieved robust and tunable protein activation in zebrafish embryos through simple addition of small molecules to the media.
- Demonstrated temporal precision in controlling enzymatic activity, including GTPase and protease function.
- Gained insights into the development of a congenital heart defect in an NRAS RASopathy mutant.
- Successfully controlled DNA and protein cleavage events catalyzed by viral recombinase and protease.
Conclusions:
- Genetic code expansion provides a powerful and broadly applicable tool for precise temporal control of protein function in zebrafish development.
- This method enables targeted chemical intervention and mechanistic studies of developmental processes and diseases.
- The developed system offers new avenues for investigating enzymatic activities and their roles in biological events.
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