Key Points

Severe heat shock enhances the immunogenicity of necrotic tumor cells.

Severe heat shock promotes cross-presentation, but not increased adjuvanticity or phagocytosis.

Enhanced cross-presentation is mediated by tumor-derived calpains.

Visual AbstractAbstract

The initiation of CD8+ T cell responses against dead cell–associated Ags is tightly regulated, facilitating adaptive immunity against pathogens and tumors while preventing autoimmunity. It is now well established that dying cells actively regulate the generation of CD8+ T cell responses via the release or exposure of damage-associated molecular patterns. However, it is unclear whether nonproteasomal proteases (activated in stressed and dying cells) can influence the availability of Ags for cross-presentation. Using a mouse model of immunogenic necrosis, we investigated the role of tumor-derived proteases in the priming of CD8+ T cells. We demonstrate that proteases released from necrotic tumor cells can degrade whole-protein Ag, generating proteolytic intermediates that are efficiently cross-presented by dendritic cells and enhance CD8+ T cell cross-priming. We identify a dominant role for calpain proteases, which are activated during necrotic cell death induced by severe heat shock. Mechanistically, proteolytic intermediates generated by tumor-derived proteases associate with necrotic tumor cell debris, which acts as a vehicle for Ag transfer that facilitates highly efficient cross-presentation in dendritic cells. Our results suggest that proteolytic systems activated in Ag donor cells during cell death may influence the availability of antigenic substrates for cross-presentation, thereby regulating the antigenicity of cell death.

Footnotes

This work was supported by an internal grant from the Division of Health Sciences, University of Otago.

The online version of this article contains supplemental material.

Abbreviations used in this article:

AFC7-amino-4-trifluoromethyl coumarinBMDCbone marrow–derived dendritic cellBODIPYboron-dipyrromethenecDC1conventional type 1 DCCRTcalreticulinDAMPdamage-associated molecular patternDCdendritic cellDPBSDulbecco’s PBSecto-CRTcell surface–exposed CRTFTfreeze-thawFTLFT lysateHSheat shockHSPheat shock proteinMFImean fluorescence intensityMHC-IMHC class IMHC-IIMHC class IImHSmild HSmHSLmHS lysateSEAPsecreted embryonic alkaline phosphatasesHSsevere HSsHSLsHS lysateReceived May 26, 2021.Accepted August 19, 2022.Copyright © 2022 by The American Association of Immunologists, Inc.