Abstract
The exposure of calreticulin (CRT) on the plasma membrane can precede anthracycline-induced apoptosis and is required for cell death to be perceived as immunogenic. Mass spectroscopy, immunofluorescence and immunoprecipitation experiments revealed that CRT co-translocates to the surface with another endoplasmic reticulum-sessile protein, the disulfide isomerase ERp57. The knockout and knockdown of CRT or ERp57 inhibited the anthracycline-induced translocation of ERp57 or CRT, respectively. CRT point mutants that fail to interact with ERp57 were unable to restore ERp57 translocation upon transfection into crt−/− cells, underscoring that a direct interaction between CRT and ERp57 is strictly required for their co-translocation to the surface. ERp57low tumor cells generated by retroviral introduction of an ERp57-specific shRNA exhibited a normal apoptotic response to anthracyclines in vitro, yet were resistant to anthracycline treatment in vivo. Moreover, ERp57low cancer cells (which failed to expose CRT) treated with anthracyclines were unable to elicit an anti-tumor response in conditions in which control cells were highly immunogenic. The failure of ERp57low cells to elicit immune responses and to respond to chemotherapy could be overcome by exogenous supply of recombinant CRT protein. These results indicate that tumors that possess an intrinsic defect in the CRT-translocating machinery become resistant to anthracycline chemotherapy due to their incapacity to elicit an anti-cancer immune response.
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Accession codes
Abbreviations
- Bac:
-
bacitracin
- CRT:
-
calreticulin
- DAPI:
-
4′,6-diamidino-2-phenylindole
- DC:
-
dendritic cells
- ER:
-
endoplasmic reticulum
- FBS:
-
fetal bovine serum
- MHC:
-
major histocompatibility complex
- PBS:
-
phosphate-buffered saline
- PDI:
-
protein disulfide isomerases
- PFA:
-
paraformaldehyde
- PI:
-
propidium iodide
- PM:
-
plasma membrane
- rec.CRT:
-
recombinant calreticulin
- recERp57:
-
recombinant ERp57
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Acknowledgements
We thank Dr Marek Michalak for the WT and crt−/− MEF and the CRT constructs. We thank Dr Natalio Garbi for the WT and erp57−/− MEF. GK is supported by the Ligue Nationale contre le Cancer (Equipe labellisée), European Commission (Active p53, Apo-Sys, RIGHT, TransDeath, ChemoRes, DeathTrain), Cancéropôle Ile-de-France, Fondation de France and Fondation pour la Recherche Médicale, TP is supported by the Swedish Research Council (Vetenskapsradet), NJ is supported by NSERC (Canada), AT is supported by Poste d'accueil INSERM and OK is supported by EMBO. K-WF and MD are grateful to Fonds zur Förderung der wissenschaftlichen Forschung (Austria) for grant ‘Molecular Enzymology’. FM is grateful to Fonds zur Förderung der wissenschaftlichen Forschung (Austria) for grant S-9304-B05. MP is partially supported by grants from Ministero della Salute and AIRC.
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Panaretakis, T., Joza, N., Modjtahedi, N. et al. The co-translocation of ERp57 and calreticulin determines the immunogenicity of cell death. Cell Death Differ 15, 1499–1509 (2008). https://doi.org/10.1038/cdd.2008.67
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DOI: https://doi.org/10.1038/cdd.2008.67
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