Deficiency of MALT1 Paracaspase Activity Results in Unbalanced Regulatory and Effector T and B Cell Responses Leading to Multiorgan Inflammation This information is current as of September 28, 2021. Frédéric Bornancin, Florian Renner, Ratiba Touil, Heiko Sic, Yeter Kolb, Ismahane Touil-Allaoui, James S. Rush, Paul A. Smith, Marc Bigaud, Ursula Junker-Walker, Christoph Burkhart, Janet Dawson, Satoru Niwa, Andreas Katopodis, Barbara Nuesslein-Hildesheim, Gisbert Weckbecker, Gerhard Zenke, Bernd Kinzel, Elisabetta Downloaded from Traggiai, Dirk Brenner, Anne Brüstle, Michael St. Paul, Natasa Zamurovic, Kathy D. McCoy, Antonius Rolink, Catherine H. Régnier, Tak W. Mak, Pamela S. Ohashi, Dhavalkumar D. Patel and Thomas Calzascia http://www.jimmunol.org/ J Immunol published online 11 March 2015 http://www.jimmunol.org/content/early/2015/03/10/jimmun ol.1402254 Supplementary http://www.jimmunol.org/content/suppl/2015/03/10/jimmunol.140225 Material 4.DCSupplemental by guest on September 28, 2021 Why The JI? Submit online. • Rapid Reviews! 30 days* from submission to initial decision • No Triage! Every submission reviewed by practicing scientists • Fast Publication! 4 weeks from acceptance to publication *average Subscription Information about subscribing to The Journal of Immunology is online at: http://jimmunol.org/subscription Permissions Submit copyright permission requests at: http://www.aai.org/About/Publications/JI/copyright.html Email Alerts Receive free email-alerts when new articles cite this article. Sign up at: http://jimmunol.org/alerts The Journal of Immunology is published twice each month by The American Association of Immunologists, Inc., 1451 Rockville Pike, Suite 650, Rockville, MD 20852 Copyright © 2015 by The American Association of Immunologists, Inc. All rights reserved. Print ISSN: 0022-1767 Online ISSN: 1550-6606. Published March 11, 2015, doi:10.4049/jimmunol.1402254 The Journal of Immunology Deficiency of MALT1 Paracaspase Activity Results in Unbalanced Regulatory and Effector T and B Cell Responses Leading to Multiorgan Inflammation Fre´de´ric Bornancin,*,1 Florian Renner,*,1 Ratiba Touil,* Heiko Sic,* Yeter Kolb,* Ismahane Touil-Allaoui,* James S. Rush,* Paul A. Smith,* Marc Bigaud,* Ursula Junker-Walker,* Christoph Burkhart,* Janet Dawson,* Satoru Niwa,* Andreas Katopodis,* Barbara Nuesslein-Hildesheim,* Gisbert Weckbecker,* Gerhard Zenke,* Bernd Kinzel,* Elisabetta Traggiai,* Dirk Brenner,†,‡ Anne Brustle,€ †,x Michael St. Paul,† Natasa Zamurovic,* Kathy D. McCoy,{ Antonius Rolink,‖ Catherine H. Re´gnier,* Tak W. Mak,† Pamela S. Ohashi,† Dhavalkumar D. Patel,* and Downloaded from Thomas Calzascia* The paracaspase MALT1 plays an important role in immune receptor-driven signaling pathways leading to NF-kB activation. MALT1 promotes signaling by acting as a scaffold, recruiting downstream signaling proteins, as well as by proteolytic cleavage of multiple substrates. However, the relative contributions of these two different activities to T and B cell function are not well http://www.jimmunol.org/ understood. To investigate how MALT1 proteolytic activity contributes to overall immune cell regulation, we generated MALT1 protease-deficient mice (Malt1PD/PD) and compared their phenotype with that of MALT1 knockout animals (Malt12/2). Malt1PD/PD mice displayed defects in multiple cell types including marginal zone B cells, B1 B cells, IL-10–producing B cells, regulatory T cells, and mature T and B cells. In general, immune defects were more pronounced in Malt12/2 animals. Both mouse lines showed abrogated B cell responses upon immunization with T-dependent and T-independent Ags. In vitro, inactivation of MALT1 protease activity caused reduced stimulation-induced T cell proliferation, impaired IL-2 and TNF-a production, as well as defective Th17 differentiation. Consequently, Malt1PD/PD mice were protected in a Th17-dependent experimental autoimmune encephalomyelitis model. Surprisingly, Malt1PD/PD animals developed a multiorgan inflammatory pathology, characterized by Th1 by guest on September 28, 2021 and Th2/0 responses and enhanced IgG1 and IgE levels, which was delayed by wild-type regulatory T cell reconstitution. We therefore propose that the pathology characterizing Malt1PD/PD animals arises from an immune imbalance featuring pathogenic Th1- and Th2/0-skewed effector responses and reduced immunosuppressive compartments. These data uncover a previously unappreciated key function of MALT1 protease activity in immune homeostasis and underline its relevance in human health and disease. The Journal of Immunology, 2015, 194: 000–000. ncontrolled NF-kB activation can lead to lymphoproli- assemble with another CARD-containing protein: CARD11 (also ferative disorders and autoimmune diseases associated known as CARMA-1), CARD9, or CARD10, respectively. The U with chronic inflammation, underscoring the need for resulting CARD-BCL10-MALT1 (CBM) complex functions as appropriate regulation of signaling via this commonly used a hub for triggering a signaling cascade that culminates in acti- pathway (1–4). The paracaspase MALT1 plays an important role vation of the IkB kinase (IKK) complex, IkB degradation, and in the regulation of NF-kB (5). Upon activation by Ag receptors, NF-kB activation (6, 7). Dectin receptors, or G protein–coupled receptors, MALT1 and the MALT1 regulates NF-kB activity in at least two ways. First, caspase recruitment domain (CARD)–containing protein BCL10 its scaffolding function is essential for NF-kB activation (8, 9). *Novartis Institutes for BioMedical Research, Novartis Pharma AG, 4056 Basel, Address correspondence and reprint requests to Dr. Thomas Calzascia, Novartis Switzerland; †Campbell Family Institute for Breast Cancer Research, Princess Margaret Institutes for BioMedical Research, Novartis Pharma AG, WSJ-386, Kohlenstrasse, Cancer Centre, University Health Network, Toronto, Ontario M5G 2C1, Canada; CH-4056 Basel, Switzerland. E-mail address: [email protected] ‡Department of Infection and Immunity, Luxembourg Institute of Health, L-4354 x The online version of this article contains supplemental material. Esch-Sur-Alzette, Luxembourg; Department of Immunology, John Curtin School of Medical Research, The Australian National University, Canberra ACT 2601, Australia; Abbreviations used in this article: BM, bone marrow; CARD, caspase recruitment {Maurice Muller€ Laboratories, University Clinic for Visceral Surgery and Medicine, domain; CBM, CARD-BCL10-MALT1 complex; cerLN, cervical and mandibular University of Bern, 3010 Bern, Switzerland; and ‖Developmental and Molecular Im- LN; CYLD, cylindromatosis; DAGG, 2,4-dinitrophenyl-b-alanyl-glycyl-glycyl-N(2- munology, Department of Biomedicine, University of Basel, 4058 Basel, Switzerland aminoethyl)carbamyl-methylated; EAE, experimental autoimmune encephalomyeli- tis; FO, follicular; GC, germinal center; IKK, IkB kinase; LN, lymph node; KLH, 1F.B. and F.R. are co-first authors. keyhole limpet hemocyanin; mesLN, mesenteric LN; MOG, myelin oligodendrocyte Received for publication September 3, 2014. Accepted for publication February 7, glycoprotein; MZ, marginal zone; PD, protease dead; PEL, peritoneal exudate cells; 2015. PP, Peyer’s patches; TCM, central memory T; TD, T-dependent; TEM, effector mem- ory T; T , follicular Th; TI, T-independent; TI-2, T-independent type 2; Treg, This work was supported by a Presidential Postdoctoral Fellowship from Novartis FH regulatory T cell; WT, wild-type. Institutes for BioMedical Research (to F.R. and H.S.) and the ATTRACT program of the Luxembourg National Research Fund (to D.B.). Copyright Ó 2015 by The American Association of Immunologists, Inc. 0022-1767/15/$25.00 www.jimmunol.org/cgi/doi/10.4049/jimmunol.1402254 2 MALT1 PROTEASE DEFICIENCY CAUSES MULTIORGAN INFLAMMATION Second, MALT1 has proteolytic activity targeting several sub- into 10-wk-old sublethally irradiated (2 times 450 rad 4 h apart) CD45.1 a b strates important in NF-kB signaling (10). These include BCL10, female WT recipients (B6.SJL-Ptprc Pepc /BoyJ; Taconic). BM cells were whose cleavage by MALT1 appears to regulate integrin-dependent isolated by flushing femur and tibiae using RPMI 1640 media and sub- sequently depleted of RBC by hypotonic shock using ACK buffer: 0.829% T cell adhesion (11). The other known substrates of MALT1 are NH4Cl, 0.1% KHCO3, and 0.0372% EDTA. all negative regulators of canonical NF-kB signaling. MALT1- dependent cleavage of the two ubiquitin editing enzymes, A20 DNP-keyhole limpet hemocyanin and 2,4-dinitrophenyl- and cylindromatosis (CYLD), downregulates their activity (12– b-alanyl-glycyl-glycyl-N(2-aminoethyl)carbamyl-methylated– 15). MALT1 also cleaves RelB, priming it for proteasome- Ficoll immunization and titer measurement dependent degradation (16) and resulting in modulation of ca- Keyhole limpet hemocyanin (KLH; Calbiochem) conjugated to DNP nonical NF-kB activation via the release of RelA and c-Rel. In (Sigma-Aldrich) was prepared, at a ratio of 1/20, as DNP-KLH stock so- addition, Regnase-1, an RNase that destabilizes a subset of NF- lution (5 mg/ml) and stored at 220˚C. DAGG (2,4-DNP-b-alanyl-glycyl- kB–dependent mRNAs by cleaving their 39 termini (e.g., c-Rel glycyl-N(2-aminoethyl)carbamyl-methylated)–Ficoll conjugate was pre- pared as described previously (21). and IL-2), is also proteolyzed and inactivated by MALT1 (17). For induction of T-dependent (TD) responses, DNP-KLH was adsorbed Thus, MALT1 regulates the NF-kB cascade by acting as a scaf- onto colloidal aluminum hydroxide (Alu-Gel-S,