The immunoproteasome, a distinct class of proteasome found predominantly in monocytes and lymphocytes, is known to shape the antigenic repertoire presented on class I major histocompatibility complexes (MHC-I). However, a specific role for the immunoproteasome in regulating other facets of immune responses has not been established. We describe here the characterization of PR-957, a selective inhibitor of low-molecular mass polypeptide-7 (LMP7, encoded by Psmb8), the chymotrypsin-like subunit of the immunoproteasome. PR-957 blocked presentation of LMP7-specific, MHC-I-restricted antigens in vitro and in vivo. Selective inhibition of LMP7 by PR-957 blocked production of interleukin-23 (IL-23) by activated monocytes and interferon-gamma and IL-2 by T cells. In mouse models of rheumatoid arthritis, PR-957 treatment reversed signs of disease and resulted in reductions in cellular infiltration, cytokine production and autoantibody levels. These studies reveal a unique role for LMP7 in controlling pathogenic immune responses and provide a therapeutic rationale for targeting LMP7 in autoimmune disorders.

Constitutive proteasomes and immunoproteasomes shape the peptide repertoire presented by major histocompatibility complex class I (MHC-I) molecules by harboring different sets of catalytically active subunits. Here, we present the crystal structures of constitutive proteasomes and immunoproteasomes from mouse in the presence and absence of the epoxyketone inhibitor PR-957 (ONX 0914) at 2.9 Å resolution. Based on our X-ray data, we propose a unique catalytic feature for the immunoproteasome subunit b5i/LMP7. Comparison of ligand-free and ligand-bound proteasomes reveals conformational changes in the S1 pocket of b5c/X but not b5i, thereby explaining the selectivity of PR-957 for b5i. Time-resolved structures of yeast proteasome:PR-957 complexes indicate that ligand docking to the active site occurs only via the reactive head group and the P1 side chain. Together, our results support structure-guided design of inhibitory lead structures selective for immunoproteasomes that are linked to cytokine production and diseases like cancer and autoimmune disorders.

Myeloid antigen-presenting cells (APC) are known to cross-present exogenous antigen on major histocompatibility class I molecules to CD8+ T cells and thereby induce protective immunity against infecting microorganisms. Here we report that liver sinusoidal endothelial cells (LSEC) are organ-resident, non-myeloid APC capable of cross-presenting soluble exogenous antigen to CD8+ T cells. Though LSEC employ similar molecular mechanisms for cross-presentation as dendritic cells, the outcome of cross-presentation by LSEC is CD8+ T cell tolerance rather than immunity. As uptake of circulating antigens into LSEC occurs efficiently in vivo, it is likely that cross-presentation by LSEC contributes to CD8+ T cell tolerance observed in situations where soluble antigen is present in the circulation.

Dendritic cells (DCs) are potent antigenpresenting cells that are able to initiate and modulate immune responses and are hence exploited as cellular vaccines for immunotherapy. Their capacity to migrate from peripheral tissues to the T-cell areas of draining lymph nodes is crucial for the priming of T lymphocytes. In this study, we investigated how the maturation of human monocyte-derived DCs (MoDCs) by several different stimuli under serum-free conditions affected their T-cell stimulatory function, cytokine secretion, and migratory behavior. Surprisingly, we found that for all maturation stimuli tested, the addition of prostaglandin E2 (PGE2) was required for effective migration of MoDCs toward the lymph nodederived chemokines CCL19 (EBI1 ligand chemokine/macrophage inflammatory protein--3␤) and CCL21 (secondary lymphoid tissue chemokine [SLC]/6Ckine). Costimulation with PGE2 enhanced the expression of the CCL19/CCL21 receptor CCR7 on the cell surface of MoDCs when they were matured with soluble CD40 ligand or proinflammatory cytokines, but did not affect CCR7 expression of polyI:C-stimulated MoDCs. The effects of PGE2 on MoDCs were mediated through increased cyclic adenosine monophosphate by 2 of the known PGE2 receptors, EP2 and EP4, which are expressed and down-regulated after PGE2 binding in these cells. In conclusion, our results suggest that signals provided by the proinflammatory mediator PGE2 are crucial for MoDCs to acquire potent T-helper cell stimulatory capacity and substantial chemotactic responsiveness to lymph node-derived chemokines. This is a new and important parameter for the preparation of IntroductionDendritic cells (DCs) are professional antigen-presenting cells that are uniquely able to stimulate naive T cells and initiate and control immune responses 1 through the activation of T-helper (T H ) cells of the T H 1, T H 2, or T H regulatory phenotype. [2][3][4] They reside in an immature stage in peripheral tissues, where they capture antigens such as invading bacteria, viruses, or damaged tissue. Upon encountering antigens, but also in response to proinflammatory cytokines or T-cell-derived signals such as CD40 ligand (CD40L), DCs lose their phagocytic capacity and up-regulate costimulatory proteins, as well as major histocompatibility complex class I and II molecules presenting processed antigen. [5][6][7] At the same time, DCs secrete large amounts of inflammatory cytokines, such as tumor necrosis factor-␣ (TNF-␣), interleukin-1␤ (IL-1␤), and IL-6, as well as chemokines (eg, CCL3 (macrophage inflammatory protein-1␣ ), CCL4 (MIP-1␤), and CCL5 (RANTES)), thus promoting the recruitment of monocytes and immature DCs into inflamed tissues. 8 Most importantly, DCs acquire the ability to migrate from peripheral tissues to the T-cell areas of draining lymphoid organs; this ability is pivotal for their capacity to initiate an immune response and is tightly regulated by several mechanisms. Migrating leukocytes express adhesion molecules, such as selectins and integrins, for the interaction with e...