Tissue-restricted inhibition of mTOR using chemical genetics
Douglas R Wassarman1,2, Kondalarao Bankapalli3, Leo J Pallanck3
1Department of Cellular and Molecular Pharmacology, University of California, San Francisco, CA 94158.
Abstract:
Mammalian target of rapamycin (mTOR) is a highly conserved eukaryotic protein kinase that coordinates cell growth and metabolism, and plays a critical role in cancer, immunity, and aging. It remains unclear how mTOR signaling in individual tissues contributes to whole-organism processes because mTOR inhibitors, like the natural product rapamycin, are administered systemically and target multiple tissues simultaneously. We developed a chemical-genetic system, termed selecTOR, that restricts the activity of a rapamycin analog to specific cell populations through targeted expression of a mutant FKBP12 protein. This analog has reduced affinity for its obligate binding partner FKBP12, which reduces its ability to inhibit mTOR in wild-type cells and tissues. Expression of the mutant FKBP12, which contains an expanded binding pocket, rescues the activity of this rapamycin analog. Using this system, we show that selective mTOR inhibition can be achieved in Saccharomyces cerevisiae and human cells, and we validate the utility of our system in an intact metazoan model organism by identifying the tissues responsible for a rapamycin-induced developmental delay in Drosophila.
Insights
Researchers developed selecTOR, a system for targeted inhibition of the mTOR pathway. This chemical-genetic tool enables studying tissue-specific mTOR signaling, crucial for understanding its role in growth, metabolism, and disease.
Area of Science:
- Molecular Biology
- Genetics
- Biochemistry
- Developmental Biology
Background:
- Mammalian target of rapamycin (mTOR) is a key regulator of cell growth, metabolism, cancer, immunity, and aging.
- Systemic administration of mTOR inhibitors like rapamycin limits the study of tissue-specific mTOR signaling roles.
- Understanding tissue-specific mTOR functions is critical for dissecting its contribution to whole-organism processes.
Purpose of the Study:
- To develop a novel chemical-genetic system for achieving cell-population-specific inhibition of mTOR signaling.
- To enable the investigation of tissue-specific roles of mTOR in complex biological processes.
- To validate the utility of the developed system in model organisms.
Main Methods:
- Development of a chemical-genetic system, selecTOR, using a rapamycin analog with reduced affinity for FKBP12.
- Targeted expression of a mutant FKBP12 protein with an expanded binding pocket to restore analog activity in specific cells.
- Validation in Saccharomyces cerevisiae, human cells, and Drosophila melanogaster to demonstrate selective mTOR inhibition.
Main Results:
- The selecTOR system successfully achieved selective mTOR inhibition in yeast and human cells.
- The system was validated in Drosophila, enabling identification of tissues mediating rapamycin-induced developmental delay.
- Demonstrated the feasibility of studying tissue-specific mTOR pathway functions using chemical genetics.
Conclusions:
- The selecTOR system provides a powerful tool for dissecting the tissue-specific functions of mTOR signaling.
- This approach overcomes limitations of systemic drug administration in studying complex biological pathways.
- Selective mTOR inhibition is achievable and valuable for understanding development and disease.
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