Abstract
Some chemotherapeutic agents can elicit apoptotic cancer cell death, thereby activating an anticancer immune response that influences therapeutic outcome. We previously reported that anthracyclins are particularly efficient in inducing immunogenic cell death, correlating with the pre-apoptotic exposure of calreticulin (CRT) on the plasma membrane surface of anthracyclin-treated tumor cells. Here, we investigated the role of cellular Ca2+ homeostasis on CRT exposure. A neuroblastoma cell line (SH-SY5Y) failed to expose CRT in response to anthracyclin treatment. This defect in CRT exposure could be overcome by the overexpression of Reticulon-1C, a manipulation that led to a decrease in the Ca2+ concentration within the endoplasmic reticulum lumen. The combination of Reticulon-1C expression and anthracyclin treatment yielded more pronounced endoplasmic reticulum Ca2+ depletion than either of the two manipulations alone. Chelation of intracellular (and endoplasmic reticulum) Ca2+, targeted expression of the ligand-binding domain of the IP3 receptor and inhibition of the sarco-endoplasmic reticulum Ca2+-ATPase pump reduced endoplasmic reticulum Ca2+ load and promoted pre-apoptotic CRT exposure on the cell surface, in SH-SY5Y and HeLa cells. These results provide evidence that endoplasmic reticulum Ca2+ levels control the exposure of CRT.
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Abbreviations
- BAPTA-AM:
-
1,2-bis(2-aminophenoxy)ethane-N,N,N′,N′-tetracetate acetyloxy methylester
- BFA:
-
brefeldin A
- BSA:
-
bovine serum albumin
- CRT:
-
calreticulin
- DAPI:
-
4′,6-diamidino-2-phenylindole
- DMEM:
-
Dulbecco's modified Eagle's medium
- ER:
-
endoplasmic reticulum
- ER-IP3R-LBD:
-
ER-targeted IP3 receptor ligand binding domain
- FBS:
-
fetal bovine serum
- KRB:
-
Krebs Ringer-modified Buffer
- mRFP:
-
monomeric Red Fluorescent Protein
- PBS:
-
phosphate-buffered Saline
- PFA:
-
paraformaldehyde
- PI:
-
propidium iodide
- PM:
-
plasma membrane
- PP1:
-
protein phosphatase 1
- RTN:
-
Reticulon
- SERCA:
-
sarcoendoplasmic reticulum Ca2+-ATPase
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Acknowledgements
We thank A Jalil for his help in confocal microscopy experiments. This work has been supported by grants from the Ligue Nationale contre le Cancer (équipe labelliseé), Institut National du Cancer (INCa), Cancéropôle Ile-de-France, the European Union (ChemoRes), ARC (Association pour la Recherche contre le Cancer), INSERM (Institut National de Recherche et Santè Medicale), and a University College London startup grant to GS.
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Tufi, R., Panaretakis, T., Bianchi, K. et al. Reduction of endoplasmic reticulum Ca2+ levels favors plasma membrane surface exposure of calreticulin. Cell Death Differ 15, 274–282 (2008). https://doi.org/10.1038/sj.cdd.4402275
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DOI: https://doi.org/10.1038/sj.cdd.4402275
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