IRAP Endosomes Control Phagosomal Maturation in Dendritic Cells

Abstract

Dendritic cells (DCs) contribute to the immune surveillance by sampling their environment through phagocytosis and endocytosis. We have previously reported that, rapidly following uptake of extracellular antigen into phagosomes or endosomes in DCs, a specialized population of storage endosomes marked by Rab14 and insulin-regulated aminopeptidase (IRAP) is recruited to the nascent antigen-containing compartment, thereby regulating its maturation and ultimately antigen cross-presentation to CD8+ T lymphocytes. Here, using IRAP–/– DCs, we explored how IRAP modulates phagosome maturation dynamics and cross-presentation. We find that in the absence of IRAP, phagosomes acquire more rapidly late endosomal markers, are more degradative, and show increased microbicidal activity. We also report evidence for a role of vesicle trafficking from the endoplasmic reticulum (ER)–Golgi intermediate compartment to endosomes for the formation or stability of the IRAP compartment. Moreover, we dissect the dual role of IRAP as a trimming peptidase and a critical constituent of endosome stability. Experiments using a protease-dead IRAP mutant and pharmacological IRAP inhibition suggest that IRAP expression but not proteolytic activity is required for the formation of storage endosomes and for DC-typical phagosome maturation, whereas proteolysis is required for fully efficient cross-presentation. These findings identify IRAP as a key factor in cross-presentation, trimming peptides to fit the major histocompatibility complex class-I binding site while preventing their destruction through premature phagosome maturation.

Mast cell-mediated inflammation relies on insulin-regulated aminopeptidase controlling cytokine export from the Golgi

“…HFI-419-treated animals secreted significantly less TNF-α and IL-6 than vehicle-treated animals, while IL-10 secretion was unaffected, indicating that the availability of the catalytic domain of IRAP was directly or indirectly required for trafficking of these pro-inflammatory cytokines. This was somewhat surprising with respect to previous studies in different cell types in which reconstitution by a protease-dead IRAP variant fully restored vesicular distribution and endosomal trafficking in IRAPko cells, suggesting that the enzymatic activity was dispensable for IRAP-mediated trafficking functions 43,45 . We therefore hypothesized that in addition to blocking its catalytic activity, HFI-419 may also affect the stability of IRAP.…”

“…Of note, in contrast to the relatively restricted Glut4 expression patterns to insulinresponsive tissues, IRAP-containing endosomes are widely expressed amongst cell types and tissues, where they are mobilized by cell-specific surface receptor signaling and employed for various cell type-specific functions (Kandror and Pilch, 2011;Bogan, 2012;Descamps et al, 2020). In this line, with regards to immune cells, the trafficking of IRAP endosomes has recently been recognized to intersect with and regulate phagosome maturation and MHC-I cross-presentation in dendritic cells (Saveanu et al, 2009;Weimershaus et al, 2012Weimershaus et al, , 2018Weimershaus et al, , 2020, activation of TLR9 (Babdor et al, 2017), as well as endo-and exocytic trafficking in T cells for the supply of TCR signaling components and optimal TCR signaling (Evnouchidou et al, 2020). Here we report that IRAP endosomes are required for the post-Golgi transport of the proinflammatory cytokines TNF-α and IL-6 in the constitutive secretion pathway in mast cells in vitro and in vivo while limiting excessive degranulation of preformed mediators.…”

“…Of note, in contrast to the relatively restricted Glut4 expression patterns to insulin-responsive tissues, IRAP-containing endosomes are widely expressed amongst cell types and tissues, where they are mobilized by cell-specific surface receptor signaling and employed for various cell type-specific functions 37–39 . In this line, with regards to immune cells, the trafficking of IRAP endosomes has recently been recognized to intersect with and regulate phagosome maturation and MHC-I cross-presentation in dendritic cells 40–43 , activation of TLR9 44 , as well as endo-and exocytic trafficking in T cells for the supply of TCR signaling components and optimal TCR signaling 45 .…”

A short ERAP2 that binds IRAP is expressed in macrophages independently from gene variation

“…IRAP instead, thanks to an additional N-terminal cytoplasmic domain, is retained in the endosomal vesicles from where it can traffic to the cell membrane forming a type II integral membrane glycoprotein (4). IRAP is a multifaceted protein: it has been shown to be involved in cross-presentation in the endosomes of dendritic cells (DC) and, when in the cell membrane, to catalyze the final step of the angiotensinogen to angiotensin IV (AT4) conversion being itself a receptor for AT4 (5,6). In addition, it has been shown to be involved in several other functions, ranging from the insulin metabolic pathway to vesicular trafficking and even in cognitive processes (7,8).…”

Discovery of Selective Inhibitor Leads by Targeting an Allosteric Site in Insulin-Regulated Aminopeptidase

“…Insulin-Regulated Aminopeptidase (IRAP, EC 3.4.11.3) is a transmembrane zinc metalloprotease that has been implicated in several important biological functions ranging from roles in glucose metabolism, cognitive functions and the immune system [ 1 , 2 , 3 , 4 ]. Currently, IRAP is under investigation as a pharmaceutical target for cognitive disorders [ 5 ].…”