Molecular dissection of plasmacytoid dendritic cell activation in vivo during a viral infection

• DOI: 10.15252/embj.201798836
• Publication Date: 2018-08-21T12:55:13Z
• Authors: Elena Tomasello; Karima Naciri; Rabie Chelbi; Gilles Bessou; Anissa Fries; Elise Gressier; Abdenour Abbas; Emeline Pollet; Philippe Pierre; Toby Lawrence; Thien‐Phong Vu Manh; Marc Dalod
• Keywords: 570; Muromegalovirus; [SDV.IMM] Life Sciences [q-bio]/Immunology; Adaptor Protein Complex 3; Interferon Regulatory Factor-7; 610; Endosomes; Microbiology; viral infection Subject Categories Immunology; Mice; 03 medical and health sciences; IRF7; Animals; Mice, Knockout; 0303 health sciences; Membrane Glycoproteins; Articles; Dendritic Cells; Herpesviridae Infections; mouse cytomegalovirus; 3. Good health; Virology & Host Pathogen Interaction; type I interferons; Toll-Like Receptor 7; plasmacytoid dendritic cells; Toll-Like Receptor 9; Interferon Type I; Myeloid Differentiation Factor 88; [SDV.IMM]Life Sciences [q-bio]/Immunology; Signal Transduction

Article21 August 2018Open Access Transparent process Molecular dissection of plasmacytoid dendritic cell activation in vivo during a viral infection Elena Tomasello Corresponding Author [email protected] Aix Marseille Univ, CNRS, INSERM, CIML, Centre d'Immunologie de Marseille-Luminy, Marseille, France Search for more papers by this author Karima Naciri Rabie Chelbi Gilles Bessou Anissa Fries Elise Gressier Abdenour Abbas Emeline Pollet Philippe Pierre orcid.org/0000-0003-0863-8255 Toby Lawrence Thien-Phong Vu Manh Marc Dalod orcid.org/0000-0002-6436-7966 Information *,1,‡, Naciri1,‡, Chelbi1,‡, Bessou1, Fries1,2, Gressier1,3, Abbas1, Pollet1, Pierre1, Lawrence1, Manh1 and *,1,‡ 1Aix 2Present address: Department Dermatology, University Hospital CHUV, Lausanne, Switzerland 3Present Microbiology Immunology, Peter Doherty Institute Infection Immunity, The Melbourne, Parkville, Vic., Australia ‡These authors contributed equally to work as senior *Corresponding author. Tel: [phone]; E-mail: 269451; EMBO Journal (2018)37:e98836https://doi.org/10.15252/embj.201798836 PDFDownload PDF article text main figures. Peer ReviewDownload summary the editorial decision including letters, reviewer comments responses feedback. ToolsAdd favoritesDownload CitationsTrack CitationsPermissions ShareFacebookTwitterLinked InMendeleyWechatReddit Figures & Info Abstract Plasmacytoid cells (pDC) are major source type I interferons (IFN-I) infections, response triggering endosomal Toll-like receptors (TLRs) 7 or 9 single-stranded RNA unmethylated CpG DNA, respectively. Synthetic ligands have been used disentangle underlying signaling pathways. adaptor protein AP3 is necessary transport molecular complexes TLRs, synthetic MyD88 into compartments allowing interferon regulatory factor (IRF7) recruitment whose phosphorylation then initiates IFN-I production. High basal expression IRF7 pDC its further enhancement positive feedback appear be robust cytokine In contrast, we show here that mouse cytomegalovirus (MCMV) produce high amounts downstream TLR9-to-MyD88-to-IRF7 pathway without requiring feedback, expression, AP3-driven routing TLRs. Hence, current model requirements professional production pDC, established using TLR ligands, does not strictly apply physiological infection. Synopsis requires both MCMV injection but function only CpG. distinct mechanisms tune TLR9 versus TLR, IFN-γ differentially contribute infection, driving largely gene programs. During injection, can independently AP3-mediated routing. low sufficient endow them with function. Cell-intrinsic LFA-1 functions hence cell-cell interactions promote their precise nature remains unknown. Introduction Type III (IFN-I/III) cytokines produced early infections playing role host resistance. Both families induce numerous interferon-stimulated genes (ISG) involved diverse biological functions, direct inhibition replication innate adaptive immunity (Tomasello et al, 2014). act on all types since receptor ubiquitously expressed, whereas IFN-III selectively targets epithelial due selective these cytokines. Upon sensing replication, through cytosolic virtually any infected IFN-I/III. However, many viruses evolved interfere detection, expressing able delay, blunt, prevent IFN-I/III cells. As likely countermeasure immune evasion strategy, vertebrates IFN-I/III-producing cells, corresponding specialized mononuclear phagocytes refractory thus escaping cell-intrinsic effects immunoevasion genes, engulf recognize material levels first identified IFN-I-producing were human (Cella 1999; Siegal 1999). well conserved along warm-blooded vertebrate evolution, based global profile (Asselin-Paturel 2001; 2002; Robbins 2008; 2015). Activated also (Yin 2012) pro-inflammatory chemokines (Swiecki Colonna, shown mice macaques (Malleret mice, resistance systemic acute herpes simplex virus 2 (HSV-2) 2013). how precisely contributes protection deciphered. shaping antiviral might supersede contribution induction IFN-I-dependent defenses, recently proposed various models 2013; Cocita 2015; Brewitz 2017). chronic macaques, humans, of, to, play protective deleterious roles tumor microenvironment inhibits polarizes toward immunosuppression, which reversed upon intratumoral administration oncolytic virus-type stimuli (Fonteneau Le Mercier several autoimmune inflammatory diseases understanding controlling specifically should help developing innovative treatments against cancer, autoimmune/inflammatory diseases. mainly activated TLR7 respective ligand pairs, uridine-containing guanosine (Shibata 2016; Zhang 2016), DNA (Kadowaki 2001), 5′-xCx (Ohto 2018). To active, TLR7/9 need UNC93B1-dependent from ER dedicated endosomes where they cleaved expose ligand-binding domain. Their MyD88-to-IRF7 leading high-level subtypes (Honda 2005a,b; Swiecki Studies Irf5-KO suggested IRF5-dependent (Takaoka 2005). This differential involvement IRF5 different has associated (Sasai 2010). cascade initiated sorted AP3-dependent pre-association IRF7. triggering, allows fast efficient NF-kB kinases IKKα IKKβ (Hoshino 2006; Pauls 2012; Hayashi most barely detectable at steady state, significantly increased via (Marie 1998; Sato 1998). constitutively express commonly accepted being key mechanism endowing (Kerkmann 2003; Honda 2005b; Prakash 2005; O'Brien 2011; Kim 2014; constitutive result tonic very homeostatic levels, vitro experiments (Taniguchi Takaoka, 2011). whether necessitates controversial. It may depend used. critical when stimulated vesicular stomatitis (VSV) (Barchet 2002) appears boost (Dalod 2002). modulation own investigated genetically deficient (Ifnar1-KO animals), abrogated enhanced perturbed responses. required rigorously investigate whether, how, modulate vivo. More generally, because studies aiming deciphering controlled conducted it crucial examine same operate address questions, combined between natural host, profiling functional ex isolated comparing results WT mutant mixed bone marrow chimera (MBMC) mice. Our canonical TLR9-to-MyD88-to-NFKB/IRF7 (CpG) administration, essential later stimulation. Thus, our study suggests trigger additional pathways compared stimulation bypass some condition. Results under distinctive instruction IFN-I, IFN-γ, Intraperitoneal leads rapid splenic TLR7/9- MyD88-dependent manner 2002, Krug 2004; Delale Zucchini 2008a,b). induces phenotypic maturation 2003). interaction mammalian broadly MyD88- Because (IFNAR) expressed types, genetic inactivation affects processes (Krug Strobl Baranek Madera Sun, Puttur 2016). generated MBMC (Fig 1A). Recipient CD45.1 lethally irradiated reconstituted 1:1 mixture (BM) Ifnar1-KO Myd88-KO CD45.2 donor animals, generate test (TST MBMC). Congenic CD45.1+ differ CD45.2+ C57BL/6 strain polymorphisms parameters (Waterstrat Therefore, included controls engrafted BM (CTR MBMC) least 8 weeks after graft. Splenic lineage− CD11b− CD11clow/int CD317high (Figs 1B EV1), 1.5 days peak production, populations 1B), pangenomic microarrays 1C–F). strategy identification was validated two independent analyses. First, showed over 94% gated SiglecH another surface marker pDC. Second, harbored transcriptional signature typical bona fide likewise ImmGen consortium EV2; Appendix Fig S1). They lacked evidence contamination other namely B T NK lymphoid cells; cDC1s, cDC2s, monocytes, macrophages, myeloid ensure transcriptomic observed strongly affected transplantation, non-irradiated animals. Figure 1. Myd88, activate sets A. Generation mice; CTR, control; TST, test. (5.1) equal proportions indicated Mice reconstitution. B. Gating strategy. lineage (TCRb, CD19, Ly6G, NK1.1)− 5.2 5.1 finally discriminated within gate. C, D. 5.2+ sorting B6 (CTR), Ifnar1−/−-, Myd88−/−-deficient (TST) 5.1+ counterparts Two performed each experimental condition (i.e., Ifnar1-CTR –TST one hand, Myd88-CTR TST hand). Samples uninfected animals empty symbols those MCMV-infected filled symbols. Altogether, led 14 replicates irrespective CD45 allotypic status originated from, MBMC. internal control. Unsupervised hierarchical clustering (HC) principal component analysis (PCA) probe microarrays. For HC, Pearson correlation distance Ward's method linkage E. Venn diagram showing overlap lists upregulated (n = 14) (gray circle), downregulated 2) (red circle) (green MBMC, 6) matched CTR F. Enrichment modules (E) previously microarray datasets (blue font), belonging specific (violet font) regulated upstream regulators (black assessed Ingenuity Pathway Analysis (IPA). Data represented heatmap, panel columns rows. colored indicate significance enrichment module set, computed IPA according color scale below heatmap. Download figure PowerPoint Click expand figure. EV1. Expression CD317 identified, 1B, (CD3, analyzed spleen (A). Dot plots represent (left panels) (right stainings representative six experiments. EV2. Comparative an compendium confirms specificity gating Principal demonstrating overall close proximity C57BL6 state conditions consortium. data batch corrected ComBat considering dataset identity cDC1, cDC2, RPM, cMo across batches, inference relationships batches (see S1 uncorrected data). Heatmap patterns well-known type-specific samples, samples (columns) (rows). Note cluster together (A) (B) individual expresses none modules. Dataset numbers correspond legend analysis, Ifnar1-TST segregated 1C). MyD88-KO Myd88-TST regrouped second branch tree. Consistently, (PCA), axis (PC1) accounting variability (26%) separated except 1D). (PC2, 17%) experiment, stronger difference than latter consistent clustering. had strong impact reprogramming, unaffected engraftment protocol much profoundly driven MyD88. identify program response, (i) 1E, gray (ii) lower (pDC IFN-I-stimulated ISG, red (iii) MyD88-stimulated MSG, green (Table EV1). We visual relative size three 1E). ISG MSG. Indeed, 56% DEGs 292) 164), 8% MSG 23). Apparently, 37% 109) neither nor induction. confirmed reprograming signaling. Representative selected heatmap illustrating 2). 2. profiles control pDCRepresentative 1E pattern Red font, (module A); D); E); black non-ISG non-MSG B). mRNA (lanes) (rows) colors above depicting classes column please refer symbol 1D. mined annotations linked pathways, diseases, regulators, ingenuity (IPA) 1F). enriched IFNB1-, IRF7-, IRF3-regulated induced CD11c+ MHC-II+ Flt3+ h subcutaneous IFN-α ("ImmGen DC ISG") (Mostafavi transcription complex IFNAR1 ISGF3, exclusive dependency exemplified OAS family Of note, among 164 dependent 50 lymphocytes emphasizing type-dependency target genes. either IFN-γ. Since no performed, apparent redundancy indirect, consecutive reciprocally. experiments, Irgm1/2 must considered archetypical targets. Unexpectedly, although Cxcl10 belonged suggesting certain known IFN-γ-stimulated such Gbp members 2), higher counterparts, antagonistic Whereas highlighted potential A D modules, former reciprocally latter. determined require vivo, delineated non-redundant shared ("DC core up") (Vu 2013) pertaining antiviral/antibacterial psoriasis. highly insulin-dependent diabetes mellitus syndrome. (D) encompassed encoding chemokines, TLR-, MyD88-, CD40-, IRAK3-, NFKB1-, IKBKB-, IRF1-, IRF3-, IRF8-regulated responses, inflammation, maturation, communication that, undergo reprogr