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Obatoclax (GX15-070) triggers necroptosis by promoting the assembly of the necrosome on autophagosomal membranes
Open accessAbstract
Obatoclax (GX15-070), a small-molecule inhibitor of antiapoptotic Bcl-2 proteins, has been reported to trigger cell death via autophagy. However, the underlying molecular mechanisms have remained elusive. Here, we identify GX15-070-stimulated assembly of the necrosome on autophagosomal membranes as a key event that connects GX15-070-stimulated autophagy to necroptosis. GX15-070 predominately induces a non-apoptotic form of cell death in rhabdomyosarcoma cells, as evident by lack of typical apoptotic features such as DNA fragmentation or caspase activation and by insensitivity to the broad-range caspase inhibitor zVAD.fmk. Instead, GX15-070 triggers massive accumulation of autophagosomes, which are required for GX15-070-induced cell death, as blockade of autophagosome formation by silencing of Atg5 or Atg7 abolishes GX15-070-mediated cell death. Co-immunoprecipitation studies reveal that GX15-070 stimulates the interaction of Atg5, a constituent of autophagosomal membranes, with components of the necrosome such as FADD, RIP1 and RIP3. This GX15-070-induced assembly of the necrosome on autophagosomes occurs in a Atg5-dependent manner, as knockdown of Atg5 abrogates formation of this complex. RIP1 is necessary for GX15-070-induced cell death, as both genetic and pharmacological inhibition of RIP1 by shRNA-mediated knockdown or by the RIP1 inhibitor necrostatin-1 blocks GX15-070-induced cell death. Similarly, RIP3 knockdown rescues GX15-070-mediated cell death and suppression of clonogenic survival. Interestingly, RIP1 or RIP3 silencing has no effect on GX15-070-stimulated autophagosome formation, underlining that RIP1 and RIP3 mediate cell death downstream of autophagy induction. Of note, GX15-070 significantly suppresses tumor growth in a RIP1-dependent manner in the chorioallantoic membrane model in vivo. In conclusion, GX15-070 triggers necroptosis by promoting the assembly of the necrosome on autophagosomes. These findings provide novel insights into the molecular mechanisms of GX15-070-induced non-apoptotic cell death.
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“necroptosis, which is favored by a necrosome-scaffolding function of the autophagy apparatus [823–825], as well as by the autophagic degradation of c-IAP1 and c-IAP2 [826].”
From Molecular mechanisms of cell death: recommendations of the nomenclature committee on cell death 2018. (2018) · cites this paper as [825]
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Metadata
- Authors
- Basit, F., Cristofanon, S., & Fulda, S.
- Affiliations
- Goethe University Frankfurt (DE)
- Corresponding author
- Fulda, S
- Venue
- Cell Death & Differentiation
- Published
- 2013-06-07
- Volume
- 20
- Issue
- 9
- Pages
- 1161-1173
- ISSN
- 1350-9047 (ISSN-L)
- 1476-5403
- PMC
- PMC3741498
- OpenAlex
- W1973671033
- Cited by
- 211
- References
- 47
- Citations by year
- 2026: 5 · 2025: 9 · 2024: 12 · 2023: 10 · 2022: 7 · 2021: 13 · 2020: 20 · 2019: 12 · 2018: 14 · 2017: 21 · 2016: 27 · 2015: 30 · 2014: 21 · 2013: 8
- FWCI
- 12.85
- Type
- journal article
- Language
- en
- Update notices
- erratum (2014-06-09)
- Open access
- bronze
- doi.org
- Locations
- Cell Death and Differentiation (publishedVersion)
- Europe PMC (PubMed Central) (publishedVersion)
- PubMed Central (publishedVersion)
- PubMed (publishedVersion)
- Funders
- Deutsche Krebshilfe
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Cell death mechanisms and regulation
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Cell Biology › Autophagy in Disease and Therapy
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Cancer Research › Ferroptosis and cancer prognosis
- Keywords
- obatoclax, necroptosis, autophagy, autophagosomes, ATG5, RIPK1, RIPK3, FADD, BCL-2 inhibitor, rhabdomyosarcoma, necrostatin-1, tumor growth inhibition, chorioallantoic membrane model, non-apoptotic cell death
- MeSH terms
- Autophagy-Related Protein 5, Autophagy-Related Protein 7, Autophagy, Cell Line, Humans, Imidazoles, Indoles, Microtubule-Associated Proteins, Necrosis, Phagosomes, Pyrroles, Cell Death, GTPase-Activating Proteins, RNA Interference, RNA, Small Interfering, Fas-Associated Death Domain Protein, Receptor-Interacting Protein Serine-Threonine Kinases, HEK293 Cells
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