Simultaneous Control of Endogenous and User-Defined Genetic Pathways Using Unique ecDHFR Pharmacological Chaperones.
Cell Chem Biol
; 27(5): 622-634.e6, 2020 05 21.
Article
in En
| MEDLINE
| ID: mdl-32330442
ABSTRACT
Destabilizing domains (DDs), such as a mutated form of Escherichia coli dihydrofolate reductase (ecDHFR), confer instability and promote protein degradation. However, when combined with small-molecule stabilizers (e.g., the antibiotic trimethoprim), DDs allow positive regulation of fusion protein abundance. Using a combinatorial screening approach, we identified and validated 17 unique 2,4-diaminopyrimidine/triazine-based ecDHFR DD stabilizers, at least 15 of which were ineffective antibiotics against E. coli and S. aureus. Identified stabilizers functioned in vivo to control an ecDHFR DD-firefly luciferase in the mouse eye and/or the liver. Next, stabilizers were leveraged to perform synergistic dual functions in vitro (HeLa cell death sensitization) and in vivo (repression of ocular inflammation) by stabilizing a user-defined ecDHFR DD while also controlling endogenous signaling pathways. Thus, these newly identified pharmacological chaperones allow for simultaneous control of compound-specific endogenous and user-defined genetic pathways, the combination of which may provide synergistic effects in complex biological scenarios.
Key words
Full text:
1
Collection:
01-internacional
Database:
MEDLINE
Main subject:
Pyrimidines
/
Tetrahydrofolate Dehydrogenase
/
Enzyme Stability
/
Folic Acid Antagonists
/
Anti-Bacterial Agents
Limits:
Animals
/
Female
/
Humans
/
Male
Language:
En
Journal:
Cell Chem Biol
Year:
2020
Document type:
Article
Affiliation country:
United States