Perforin-2 is a pore-forming effector of endocytic escape in cross-presenting dendritic cells

Abstract

During initiation of antiviral and antitumor T cell–mediated immune responses, dendritic cells (DCs) cross-present exogenous antigens on major histocompatibility complex (MHC) class I molecules. Cross-presentation relies on the unusual “leakiness” of endocytic compartments in DCs, whereby internalized proteins escape into the cytosol for proteasome-mediated generation of MHC I–binding peptides. Given that type 1 conventional DCs excel at cross-presentation, we searched for cell type–specific effectors of endocytic escape. We devised an assay suitable for genetic screening and identified a pore-forming protein, perforin-2 ( Mpeg1 ), as a dedicated effector exclusive to cross-presenting cells. Perforin-2 was recruited to antigen-containing compartments, where it underwent maturation, releasing its pore-forming domain. Mpeg1 −/− mice failed to efficiently prime CD8 + T cells to cell-associated antigens, revealing an important role for perforin-2 in cytosolic entry of antigens during cross-presentation.

HOPS-Dependent Endosomal Escape Demands Protein Unfolding

“…The observation that ZF5.3-mediated endosomal escape is most efficient when conjugated to low- T m proteins, and that this pathway demands communication between luminal ZF5.3 and cytosol-facing HOPS, suggests the existence of a selective portal through which membrane transport occurs. In nearly all cases, nature mediates such transport via a proteinaceous channel embedded within the membrane, such as the recently reported perforin-2 channel in dendritic cells . Whether ZF5.3 accomplishes its escape via lipid interactions or makes use of a yet-undetected protein channel remains under active investigation.…”

“…In nearly all cases, nature mediates such transport via a proteinaceous channel embedded within the membrane, such as the recently reported perforin-2 channel in dendritic cells. 59 Whether ZF5.3 accomplishes its escape via lipid interactions or makes use of a yet-undetected protein channel remains under active investigation. Regardless, the results of this study provide clear biophysical guidelines to promote endosomal escape of ZF5.3-tagged cargo and can be applied to the development and expansion of novel protein therapies.…”

Tim4 enables large peritoneal macrophages to cross-present tumor antigens at early stages of tumorigenesis

“…First, receptors that mediate the uptake of dead cells such as Clec9-DNGR-1, CD36, Axl, SCARF-1 and TIM3 have been suggested to drive cross-presentation of ingested antigens in dendritic cells 42–47 . Second, depending on the cellular context, cross-presentation is facilitated by decreased acidification in phagosomes to preserve antigen integrity; active transfer to the cytosol to intersect the conventional MHC class-I pathway and specialized pathways of vacuolar trafficking 42,48–51 .…”

Cytotoxicity as a form of immunogenic cell death leading to efficient tumor antigen cross‐priming

“…18 More recently, perforin-2 was shown to facilitate the endosomal delivery of antigens to the cytosol in cross-presenting dendritic cells, leading to the impairment of cross-priming and CD8 + T cell responses in vivo. 24 All of these mouse models have provided crucial insights into the mechanisms underlying antigen crosspresentation and T cell cross-priming, emphasizing not only the pivotal role of cDC1s in initiating and maintaining potent antitumor immune responses but also highlighting that cross-priming is one of the most important functions exerted by cDC1 cells in cancer.…”