Abstract
Mutations resulting in decreased activity of p53 tumor suppressor protein promote tumorigenesis. P53 protein levels are tightly regulated through the Ubiquitin Proteasome System (UPS). Several E3 ligases were shown to regulate p53 stability, including MDM2. Here we report that the ubiquitin E3 ligase XIAP (X-linked Inhibitors of Apoptosis) is a direct ligase for p53 and describe a novel approach for modulating the levels of p53 by targeting the XIAP pathway. Using in vivo (live-cell) and in vitro (cell-free reconstituted system) ubiquitylation assays, we show that the XIAP-antagonist ARTS regulates the levels of p53 by promoting the degradation of XIAP. XIAP directly binds and ubiquitylates p53. In apoptotic cells, ARTS inhibits the ubiquitylation of p53 by antagonizing XIAP. XIAP knockout MEFs express higher p53 protein levels compared to wild-type MEFs. Computational screen for small molecules with high affinity to the ARTS-binding site within XIAP identified a small-molecule ARTS-mimetic, B3. This compound stimulates apoptosis in a wide range of cancer cells but not normal PBMC (Peripheral Blood Mononuclear Cells). Like ARTS, the B3 compound binds to XIAP and promotes its degradation via the UPS. B3 binding to XIAP stabilizes p53 by disrupting its interaction with XIAP. These results reveal a novel mechanism by which ARTS and p53 regulate each other through an amplification loop to promote apoptosis. Finally, these data suggest that targeting the ARTS binding pocket in XIAP can be used to increase p53 levels as a new strategy for developing anti-cancer therapeutics.
| Original language | English |
|---|---|
| Pages (from-to) | 1145-1160 |
| Number of pages | 16 |
| Journal | Apoptosis |
| Volume | 29 |
| Issue number | 7-8 |
| DOIs | |
| Publication status | Published - 29 Apr 2024 |
Bibliographical note
We thank Prof. Hermann Steller for thoughtful input and discussions of the manuscript. We thank Dr. Meirav Avita-Shacham, head of the analytical and chromatography unit, Dr. Boris Shklyar, head of the Bio-imaging Unit, and Dr. Sagie Schif, head of the flow Cytometry Unit, at the University of Haifa. We also thank Dr. Maya Lalzar, head of Bioinformatics Services Unit, Faculty of Natural Sciences, University of Haifa, for her thorough proteome analysis, and Dr. Moran Jerabek for performing and analyzing the MST binding assays (at Crelux a WuXi AppTech company Ltd, Germany). We also thank Dr. Donald Alastair for his work on the in silico screen for synthetic small molecules that bind to the specific ARTS binding site within XIAP-BIR3 (at BioSolveIT). This work was funded by Israel Science Foundation (ISF) Grant 822/12 (to S.L.) and by a generous grant award from the Hymen Milgrom Trust (to S.L.). Also, funded by the U.S. Israel Binational Science Foundation Grant 2003085, Israel Science Foundation (ISF) Grants 1264/06, INCPM-ISF Grant 2376/15.Open access funding provided by University of Haifa. This work was funded by Israel Science Foundation (ISF) Grant 822/12 (to S.L.) and by a generous grant award from the Hymen Milgrom Trust (to S.L.). Also, funded by the U.S. Israel Binational Science Foundation Grant 2003085, Israel Science Foundation (ISF) Grants 1264/06, INCPM-ISF Grant 2376/15.
Publisher Copyright:
© The Author(s) 2024.
All Science Journal Classification (ASJC) codes
- Pharmacology
- Pharmaceutical Science
- Clinical Biochemistry
- Cell Biology
- Biochemistry, medical
- Cancer Research