title: Immunoproteasome dysfunction augments alternative polarization of alveolar macrophages.
abstract: The proteasome is a central regulatory hub for intracellular signaling by degrading numerous signaling mediators. Immunoproteasomes are specialized types of proteasomes involved in shaping adaptive immune responses, but their role in innate immune signaling is still elusive. Here, we analyzed immunoproteasome function for polarization of alveolar macrophages, highly specialized tissue macrophages of the alveolar lung surface. Classical activation (M1 polarization) of primary alveolar macrophages by LPS/IFNγ transcriptionally induced all three immunoproteasome subunits, low molecular mass protein 2 (LMP2), LMP7 and multicatalytic endopeptidase complex-like 1, which was accompanied by increased immunoproteasome activity in M1 cells. Deficiency of LMP7 had no effect on the LPS/IFNγ-triggered M1 profile indicating that immunoproteasome function is dispensable for classical alveolar macrophage activation. In contrast, IL-4 triggered alternative (M2) activation of primary alveolar macrophages was accompanied by a transcriptionally independent amplified expression of LMP2 and LMP7 and an increase in immunoproteasome activity. Alveolar macrophages from LMP7 knockout mice disclosed a distorted M2 profile upon IL-4 stimulation as characterized by increased M2 marker gene expression and CCL17 cytokine release. Comparative transcriptome analysis revealed enrichment of IL-4-responsive genes and of genes involved in cellular response to defense, wounding and inflammation in LMP7-deficient alveolar macrophages indicating a distinct M2 inflammation resolving phenotype. Moreover, augmented M2 polarization was accompanied by amplified AKT/STAT6 activation and increased RNA and protein expression of the M2 master transcription factor interferon regulatory factor 4 in LMP7(-/-) alveolar macrophages. IL-13 stimulation of LMP7-deficient macrophages induced a similar M2-skewed profile indicative for augmented signaling via the IL-4 receptor α (IL4Rα). IL4Rα expression was generally elevated only on protein but not RNA level in LMP7(-/-) alveolar macrophages. Importantly, specific catalytic inhibition with an LMP7-specific proteasome inhibitor confirmed augmented IL-4-mediated M2 polarization of alveolar macrophages. Our results thus suggest a novel role of immunoproteasome function for regulating alternative activation of macrophages by limiting IL4Rα expression and signaling.
authors.affiliation: Comprehensive Pneumology Center (CPC), Institute of Lung Biology and Disease, Helmholtz Zentrum München, Neuherberg, Germany.|Department of Pathophysiology, West China School of Preclinical Sciences and Forensic Medicine, Sichuan University, Chengdu, Sichuan Province, China.
authors.forename: S
authors.initials: S
authors.lastname: Chen
authors.affiliation: Comprehensive Pneumology Center (CPC), University Hospital, Ludwig-Maximilians University and Helmholtz Zentrum München, Member of the German Center for Lung Research (DZL), Munich, Germany.
authors.forename: I E
authors.initials: IE
authors.lastname: Kammerl
authors.affiliation: Comprehensive Pneumology Center (CPC), University Hospital, Ludwig-Maximilians University and Helmholtz Zentrum München, Member of the German Center for Lung Research (DZL), Munich, Germany.
authors.forename: O
authors.initials: O
authors.lastname: Vosyka
authors.affiliation: Comprehensive Pneumology Center (CPC), University Hospital, Ludwig-Maximilians University and Helmholtz Zentrum München, Member of the German Center for Lung Research (DZL), Munich, Germany.
authors.forename: T
authors.initials: T
authors.lastname: Baumann
authors.affiliation: Comprehensive Pneumology Center (CPC), Institute of Lung Biology and Disease, Helmholtz Zentrum München, Neuherberg, Germany.
authors.forename: Y
authors.initials: Y
authors.lastname: Yu
authors.affiliation: Max von Pettenkofer-Institute, Ludwig-Maximilians University, Munich, Germany.
authors.forename: Y
authors.initials: Y
authors.lastname: Wu
authors.affiliation: Institute of Experimental Genetics, Helmholtz Zentrum München, Neuherberg, Germany.
authors.forename: M
authors.initials: M
authors.lastname: Irmler
authors.affiliation: Department of Bio-Organic Synthesis, Leiden University, Leiden, The Netherlands.
authors.forename: H S
authors.initials: HS
authors.lastname: Overkleeft
authors.affiliation: Institute of Experimental Genetics, Helmholtz Zentrum München GmbH, Neuherberg, Germany.|German Center for Diabetes Research (DZD e.V.), Neuherberg, Germany.|Technische Universität München, Chair of Experimental Genetics, Freising, Germany.
authors.forename: J
authors.initials: J
authors.lastname: Beckers
authors.affiliation: Comprehensive Pneumology Center (CPC), Institute of Lung Biology and Disease, Helmholtz Zentrum München, Neuherberg, Germany.|Comprehensive Pneumology Center (CPC), University Hospital, Ludwig-Maximilians University and Helmholtz Zentrum München, Member of the German Center for Lung Research (DZL), Munich, Germany.
authors.forename: O
authors.initials: O
authors.lastname: Eickelberg
authors.affiliation: Comprehensive Pneumology Center (CPC), University Hospital, Ludwig-Maximilians University and Helmholtz Zentrum München, Member of the German Center for Lung Research (DZL), Munich, Germany.
authors.forename: S
authors.initials: S
authors.lastname: Meiners
authors.affiliation: Comprehensive Pneumology Center (CPC), Institute of Lung Biology and Disease, Helmholtz Zentrum München, Neuherberg, Germany.
authors.forename: T
authors.initials: T
authors.lastname: Stoeger
mesh_terms: D000818:Animals; D016764:Cell Polarity / Q000187:drug effects; D002478:Cells, Cultured; D054414:Chemokine CCL17 / Q000032:analysis; D003546:Cysteine Endopeptidases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D050835:Interferon Regulatory Factors / Q000378:metabolism; D007371:Interferon-gamma / Q000494:pharmacology; D018793:Interleukin-13 / Q000494:pharmacology; D015847:Interleukin-4 / Q000032:analysis; D008070:Lipopolysaccharides / Q000633:toxicity; D016676:Macrophages, Alveolar / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D051379:Mice; D008807:Mice, Inbred BALB C; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D009842:Oligopeptides / Q000494:pharmacology; D046988:Proteasome Endopeptidase Complex / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D015854:Up-Regulation / Q000187:drug effects; D000099218:Interferon Regulatory Factor-4
chemical_list: D054414:Chemokine CCL17; D003546:Cysteine Endopeptidases; D050835:Interferon Regulatory Factors; D007371:Interferon-gamma; D018793:Interleukin-13; D015847:Interleukin-4; D008070:Lipopolysaccharides; D009842:Oligopeptides; D046988:Proteasome Endopeptidase Complex; D051057:Proto-Oncogene Proteins c-akt; D000099218:Interferon Regulatory Factor-4; C542291:PR-957; C071313:LMP-2 protein; C077436:LMP7 protein