TY - JOUR
T1 - A novel auxin-inducible degron system for rapid, cell cycle-specific targeted proteolysis
AU - Capece, Marina
AU - Tessari, Anna
AU - Mills, Joseph
AU - Vinciguerra, Gian Luca Rampioni
AU - Louke, Darian
AU - Lin, Chenyu
AU - McElwain, Bryan K.
AU - Miles, Wayne O.
AU - Coppola, Vincenzo
AU - Davies, Alexander E.
AU - Palmieri, Dario
AU - Croce, Carlo M.
N1 - Publisher Copyright:
© 2023, The Author(s), under exclusive licence to ADMC Associazione Differenziamento e Morte Cellulare.
PY - 2023/9
Y1 - 2023/9
N2 - The discrimination of protein biological functions in different phases of the cell cycle is limited by the lack of experimental approaches that do not require pre-treatment with compounds affecting the cell cycle progression. Therefore, potential cycle-specific biological functions of a protein of interest could be biased by the effects of cell treatments. The OsTIR1/auxin-inducible degron (AID) system allows “on demand” selective and reversible protein degradation upon exposure to the phytohormone auxin. In the current format, this technology does not allow to study the effect of acute protein depletion selectively in one phase of the cell cycle, as auxin similarly affects all the treated cells irrespectively of their proliferation status. Therefore, the AID system requires coupling with cell synchronization techniques, which can alter the basal biological status of the studied cell population, as with previously available approaches. Here, we introduce a new AID system to Regulate OsTIR1 Levels based on the Cell Cycle Status (ROLECCS system), which induces proteolysis of both exogenously transfected and endogenous gene-edited targets in specific phases of the cell cycle. We validated the ROLECCS technology by down regulating the protein levels of TP53, one of the most studied tumor suppressor genes, with a widely known role in cell cycle progression. By using our novel tool, we observed that TP53 degradation is associated with increased number of micronuclei, and this phenotype is specifically achieved when TP53 is lost in S/G2/M phases of the cell cycle, but not in G1. Therefore, we propose the use of the ROLECCS system as a new improved way of studying the differential roles that target proteins may have in specific phases of the cell cycle.
AB - The discrimination of protein biological functions in different phases of the cell cycle is limited by the lack of experimental approaches that do not require pre-treatment with compounds affecting the cell cycle progression. Therefore, potential cycle-specific biological functions of a protein of interest could be biased by the effects of cell treatments. The OsTIR1/auxin-inducible degron (AID) system allows “on demand” selective and reversible protein degradation upon exposure to the phytohormone auxin. In the current format, this technology does not allow to study the effect of acute protein depletion selectively in one phase of the cell cycle, as auxin similarly affects all the treated cells irrespectively of their proliferation status. Therefore, the AID system requires coupling with cell synchronization techniques, which can alter the basal biological status of the studied cell population, as with previously available approaches. Here, we introduce a new AID system to Regulate OsTIR1 Levels based on the Cell Cycle Status (ROLECCS system), which induces proteolysis of both exogenously transfected and endogenous gene-edited targets in specific phases of the cell cycle. We validated the ROLECCS technology by down regulating the protein levels of TP53, one of the most studied tumor suppressor genes, with a widely known role in cell cycle progression. By using our novel tool, we observed that TP53 degradation is associated with increased number of micronuclei, and this phenotype is specifically achieved when TP53 is lost in S/G2/M phases of the cell cycle, but not in G1. Therefore, we propose the use of the ROLECCS system as a new improved way of studying the differential roles that target proteins may have in specific phases of the cell cycle.
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U2 - 10.1038/s41418-023-01191-4
DO - 10.1038/s41418-023-01191-4
M3 - Article
C2 - 37537305
AN - SCOPUS:85166664245
SN - 1350-9047
VL - 30
SP - 2078
EP - 2091
JO - Cell Death and Differentiation
JF - Cell Death and Differentiation
IS - 9
ER -