Mitarbeiter:innen

Prof. Marcus Altfeld, Benjamin Grünhagel, Dr. Maria Pujantell Graell, Rebecco-Jo Thiele, Dr. Kati Tillack

Forschungsprojekte

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Our immune system defends us from environmental threats, such as bacterial and viral infections, and detects and removes abnormal cells that potentially lead to malignancies. Optimal immunological homeostasis is achieved when the threat (e.g. pathogen) is removed with the highest efficiency whilst avoiding collateral tissue damage for the host. More and more scientific data, however, suggest that this immunological balance differs between women and men. It has been shown that women mount stronger immune responses against pathogens compared to men, leading to more rapid control or clearance of infections. Women also develop stronger immunity in response to vaccinations, and exhibit stronger immune responses against malignancies, including leukemia. Enhanced immune responsiveness in females, however, comes at a cost, including aggravated tissue damage, higher immune activation, persistent inflammation and significantly higher incidences of autoimmune diseases, such as autoimmune hepatitis and multiple sclerosis. However, the precise contribution of sex hormones and sex chromosome-encoded genes to sexspecific differences in immunity, and their relevance for human disease manifestations, remains insufficiently understood. The understanding of these mechanisms will pave the way for novel treatment strategies for infectious and immune-mediated diseases that take the sex of the affected individual into account.

FOR 5068 "Sex differences in Immunity"

Type I IFN-mediated sex differences in immune response to HIV-1

In acute HIV-1 infection, women control viral replication better than men. In contrast, women develop increased immune activation and faster loss of CD4+ T cells during untreated chronic HIV-1 infection. Increasing data indicate that these sex differences in the manifestation of HIV-1 disease are mediated by sex-specific differences in antiviral immunity. In previous studies we demonstrated that pDCs derived from women produce more IFNα in response to HIV-1 than pDCs from men, potentially contributing to the better control of viremia in acute infection. This increased IFNα production in response to HIV-1 however also led to significantly higher immune activation and faster loss of CD4+ T cells in chronically HIV-1-infected women. Taken together these data strongly suggest that sex differences in Type I IFN production result in differences in HIV-1 disease manifestations between women and men. However, the precise molecular mechanisms underlying these sex differences in HIV-1 and other infectious diseases remain poorly understood. We hypothesize that X chromosome-encoded genes play a critical role in regulating Type I IFN responses, and that gene-dose effects resulting from escape from X chromosome inactivation (XCI) contribute to sex-specific differences in antiviral immunity, using HIV-1 as a model. We will furthermore test the hypothesis that changes in levels of sex hormones can regulate Type I IFN responses of pDCs, using longitudinal samples from a human transgender cohort. Taken together, these studies will identify critical mechanisms underlying sex-specific differences in antiviral immunity, and provide rationale for the design of interventions that take these differences between women and men into account.

Forschungsnetzwerke

DFG: KFO 306 Primär Sklerosierende Cholangitis

DFG: SFB 841 Leberentzündung: Infektion, Regulation und Konsequenzen (A7: Mechanismen der NK-Zell-vermittelten Kontrolle der HCV-Infektion)

EHVA: European HIV Alliance

DZIF: Deutsches Zentrum für Infektionsforschung

Ausgewählte Publikationen

Hagen SH, Hennesen J, Altfeld M. (2021) Assessment of escape from X chromosome inactivation and gene expression in single human immune cells. STAR Protoc 2:100641

Sven Hendrik Hagen, Florian Henseling, Jana Hennesen, Hélène Savel, Solenne Delahaye, Laura Richert, Susanne Maria Ziegler, Marcus Altfeld. (2020) Heterogeneous Escape from X Chromosome Inactivation Results in Sex Differences in Type I IFN Responses at the Single Human pDC Level. Cell Rep 33:108485

Adland E, Millar J, Bengu N, Muenchhoff M, Fillis R, Sprenger K, Ntlantsana V, Roider J, Vieira V, Govender K, Adamson J, Nxele N, Ochsenbauer C, Kappes J, Mori L, van Lobenstein J, Graza Y, Chinniah K, Kapongo C, Bhoola R, Krishna M, Matthews PC, Poderos RP, Lluch MC, Puertas MC, Prado JG, McKerrow N, Archary M, Ndung'u T, Groll A, Jooste P, Martinez-Picado J, Altfeld M, Goulder P. (2020) Sex-specific innate immune selection of HIV-1 in utero is associated with increased female susceptibility to infection. Nat Commun 11:1767

Bunders MJ, Altfeld M (2020) Implications of Sex Differences in Immunity for SARS-CoV-2 Pathogenesis and Design of Therapeutic Interventions. Immunity 53:487-495

Lotter H, Altfeld M. (2019) Sex differences in immunity. Semin Immunopathol. 41:133-135

Scully EP, Gandhi M, Johnston R, Hoh R, Lockhart A, Dobrowolski C, Pagliuzza A, Milush JM, Baker CA, Girling V, Ellefson A, Gorelick R, Lifson J, Altfeld M, Alter G, Cedars M, Solomon A, Lewin SR, Karn J, Chomont N, Bacchetti P, Deeks SG. (2019) Sex-Based Differences in Human Immunodeficiency Virus Type 1 Reservoir Activity and Residual Immune Activation. J Infect Dis. 219:1084-1094

Rechtien A, Altfeld M. (2019) Sexual dimorphism in HIV-1 infection. Semin Immunopathol 41:195-202

Ziegler SM, Beisel C, Sutter K, Griesbeck M, Hildebrandt H, Hagen SH, Dittmer U, Altfeld M. (2017) Human pDCs display sex-specific differences in type I interferon subtypes and interferon α/β receptor expression. Eur J Immunol 47:251-256

Griesbeck M, Ziegler S, Laffont S, Smith N, Chauveau L, Tomezsko P, Sharei A, Kourjian G, Porichis F, Hart M, Palmer CD, Sirignano M, Beisel C, Hildebrandt H, Cénac C, Villani AC, Diefenbach TJ, Le Gall S, Schwartz O, Herbeuval JP, Autran B, Guéry JC, Chang JJ, Altfeld M. (2015) Sex Differences in Plasmacytoid Dendritic Cell Levels of IRF5 Drive Higher IFN-α Production in Women. J Immunol. 195:5327-36

Addo MM, Altfeld M. (2014) Sex-based differences in HIV type 1 pathogenesis. J Infect Dis. 209 Suppl 3:S86-92

Chang JJ, Woods M, Lindsay RJ, Doyle EH, Griesbeck M, Chan ES, Robbins GK, Bosch RJ, Altfeld M. (2013) Higher expression of several interferon-stimulated genes in HIV-1-infected females after adjusting for the level of viral replication. J Infect Dis 208:830-8

Meier A, Chang JJ, Chan ES, Pollard RB, Sidhu HK, Kulkarni S, Wen TF, Lindsay RJ, Orellana L, Mildvan D, Bazner S, Streeck H, Alter G, Lifson JD, Carrington M, Bosch RJ, Robbins GK, Altfeld M. (2009) Sex differences in the Toll-like receptor-mediated response of plasmacytoid dendritic cells to HIV-1. Nat Med 15: 955-9

Mitarbeiter:innen

Prof. Friedrich Haag, Gudrun Dubberke, Samantha Eiberg, Kilian Keetz, Julia Radko

Forschungsprojekte

Die Bedeutung extrazellulärer Adenin-Nukleotide für die Regulation der Immunantwort bei Virusinfektionen und Tumorerkrankungen

Die Immunantwort gegen eine Virusinfektion ist immer mit Gewebsschädigung verbunden, da die Immunreaktion auf die Zerstörung infizierter Zellen - in der Regel durch zytotoxische T Lymphozyten - abzielt. Die Krankheitssymptomatik ergibt sich daher aus der Balance zwischen der direkten gewebsschädigenden Wirkung des Virus und der Schädigung durch den Angriff des Immunsystems. In manchen Fällen, wie z.B. bei schweren Krankheitsverläufen der SARS-Cov-2 Infektion, verursacht die überschießende Immunreaktion einen größeren Schaden als die direkte Virus-bedingte Zellschädigung.

Das Vorkommen der Adenin-Nukleotide ATP (Adenosintriphosphat) und Adenosin in der Gewebsflüssigkeit im entzündeten Gewebe hat einen großen Einfluss auf die Stärke und Ausprägung der Immunantwort. ATP wird von geschädigten Zellen freigesetzt und wirkt durch Bindung an P2X Rezeptoren entzündungsverstärkend. Gleichzeitig wird ATP durch die Oberflächenenzyme CD39 und CD73 zu Adenosin abgebaut, welches seinerseits durch Bindung an P1 Rezeptoren die Funktionen vieler Immunzellen dämpft.

Unsere Arbeitsgruppe untersucht die Verteilung von CD39 und CD73 auf der Oberfläche unterschiedlicher Populationen von Virus-spezifischen T-Lymphozyten. Insbesondere interessiert uns dabei, wie ATP und ADO die Funktionen dieser Zellen, d.h. ihre Zytotoxizität und ihre Fähigkeit, Botenstoffe (Zytokine) zu sezernieren beeinflussen. Ebenfalls interessieren uns mögliche Wechselwirkungen zwischen der Immunregulation durch Adenin-Nukleotide und der T-Zell Erschöpfung, einem weiteren Mechanismus zur Begrenzung von Gewebsschäden im Rahmen chronischer Virusinfektionen, der durch das Vorkommen hemmender Rezeptoren auf der Oberfläche Virus-spezifischer T-Zellen gekennzeichnet ist.

Verwendung fluoreszierender ATP-Sensoren zur bildlichen Darstellung von extrazellulärem ATP

Die Ausschleusung von ATP liefert den Rohstoff für alle weiteren Schritte der Regulation von Immunfunktionen durch extrazelluläre Adenin-Nukleotide. Um die Bedeutung dieses Mechanismus richtig einschätzen zu können ist es wichtig zu wissen, in welcher Konzentration ATP an den Rezeptoren auf Zelloberflächen ankommt und wie schnell es wieder abgebaut wird. Um diese Frage zu untersuchen testen wir Moleküle, die durch Bindung von ATP ihre Fluoreszenzeigenschaften verändern, auf ihre Eignung als ATP-Sensoren in bildgebenden Verfahren wie der Lebendzell-Mikroskopie. Durch Einbringen der Gensequenz für einen solchen Sensor, so dass er entweder im Inneren von Zellen (Zytoplasma) oder an ihrer Oberfläche gemacht wird, konnten wir zeigen, dass Aktivierung des P2X7 Rezeptors zu einer Abnahme des ATP-Gehalts im Inneren der Zelle bei gleichzeitiger Zunahme an der Zelloberfläche führt (Abb 2).

Gleichzeitig arbeiten wir an der Produktion von löslichen ATP-Sensoren, die wir von außen an Zellmembranen anbringen können, um den Austritt von ATP besser visualisieren zu können. Dabei handelt es sich entweder um fluoreszierende, ATP-bindende Proteine, welche gentechnisch hergestellt werden, oder, in Zusammenarbeit mit der Arbeitsgruppe von Chris Meier am Institut für Organische Chemie der Universität Hamburg, um kleine Moleküle. Dabei ist es eine besondere Herausforderung, die Sensoren so zu verändern, dass sie gezielt an Zelloberflächen gebracht werden können.

Verfolgen der SARS-Cov2-spezifischen T-Zell Antwort bei immunsupprimierten Personen nach Infektion oder Immunisierung

Die Impfung gegen das Spike Protein des SARS-Cov2 Virus erzeugt neben neutralisierenden Antikörpern, die vor einer Infektion mit dem Virus schützen, auch eine T-Zell-basierte Immunantwort, die zum Schutz vor einem schweren Krankheitsverlauf beitragen. Patienten unter immunsupprimierender Medikation, z.B. infolge einer Organtransplantation, stellen eine besonders vulnerable Gruppe gegenüber einer Infektion mit dem SARS-Cov2 Virus dar. Bei einem Teil dieser Patienten erzeugt die Immunisierung nur einen geringen Infektionsschutz in Form neutralisierender Antikörper. Um die Stärke und Verlauf der T-Zell-basierten Immunantwort über die Zeit verfolgen zu können, haben wir Methoden entwickelt, um die spezifische T-Zell Antwort gegen SARS-Cov2 Antigene zu erfassen. Die Ergebnisse zeigen, dass immunsupprimierte Patienten sehr unterschiedlich reagieren und dass eine erfolgreiche Antikörperantwort nicht immer mit einer starken T-Zell-Antwort einhergeht und umgekehrt. Sie legen nahe, dass bei Patienten unter immunsuppressiver Therapie die Immunantwort auf die Impfung nachverfolgt werden sollte, um optimale Zeitpunkte für eine Auffrischimpfung zu ermitteln.

Forschungsnetzwerke

DFG: SFB 1328 Adenine nucleotides in immunity and inflammation (A12 Role of the ectonucleotidases CD39 and CD73 for the modulation of the immune response in chronic human viral infections)

DFG: SFB 1328 Adenine nucleotides in immunity and inflammation (Z02 Antibodies and nanobodies for adenine nucleotide research)

Ausgewählte Publikationen

REGULATION VON IMMUNFUNKTIONEN DURCH EXTRAZELLULÄRE NUKLEOTIDE

Brauneck F, Haag F, Woost R, Wildner N, Tolosa E, Rissiek A, Vohwinkel G, Wellbrock J, Bokemeyer C, Schulze zur Wiesch J, Ackermann C, and Fiedler W. (2021) Increased frequency of TIGIT+CD73-CD8+ T cells with a TOX+ TCF-1low profile in patients with newly diagnosed and relapsed AML. OncoImmunology. 10:193039110.1080/2162402X.2021.1930391.

Libera J, Wittner M, Kantowski M, Woost R, Eberhard JM, de Heer J, Reher D, Huber S, Haag F, and Schulze Zur Wiesch J. (2020) Decreased Frequency of Intestinal CD39(+) γδ(+) T Cells With Tissue-Resident Memory Phenotype in Inflammatory Bowel Disease. Front Immunol. 11:56747210.3389/fimmu.2020.567472.

Haag F, and Buck F. (2015) Identification and analysis of ADP-ribosylated proteins. Curr Top Microbiol Immunol. 384:33-5010.1007/82_2014_424.

Seman M, Adriouch S, Scheuplein F, Krebs C, Freese D, Glowacki G, Deterre P, Haag F, and Koch-Nolte F. (2003) NAD-induced T cell death: ADP-ribosylation of cell surface proteins by ART2 activates the cytolytic P2X7 purinoceptor. Immunity. 19:571-58210.1016/s1074-7613(03)00266-8.

ATP SENSOREN

Kaschubowski KE, Kraft AE, Nikolaev VO, and Haag F. (2020) Using FRET-Based Fluorescent Sensors to Monitor Cytosolic and Membrane-Proximal Extracellular ATP Levels. Methods Mol Biol. 2041:223-23110.1007/978-1-4939-9717-6_16.

Johnsen B, Kaschubowski KE, Nader S, Schneider E, Nicola JA, Fliegert R, Wolf IMA, Guse AH, Nikolaev VO, Koch-Nolte F, and Haag F. (2019) P2X7-mediated ATP secretion is accompanied by depletion of cytosolic ATP. Purinergic Signal. 15:155-16610.1007/s11302-019-09654-5.

DIE IMMUNANTWORT BEI INFEKTION UND IMMUNISIERUNG GEGEN SARS-COV-2

Haag F. (2021) Die natürliche und adaptive Immunantwort bei der Covid-19 Erkrankung Hamburger Ärzteblatt. 75:28-30

Rüther DF, Schaub GM, Duengelhoef PM, Haag F, Brehm TT, Fathi A, Wehmeyer M, Jahnke-Triankowski J, Mayer L, Hoffmann A, Fischer L, Addo MM, Luetgehetmann M, Lohse AW, Schulze Zur Wiesch J, and Sterneck M. (2021) SARS-CoV2-specific humoral and T-cell immune response after second vaccination in liver cirrhosis and transplant patients. Clinical Gastroenterology and Hepatology. https://doi.org/10.1016/j.cgh.2021.09.003.

Woo MS, Haag F, Nierhaus A, Jarczak D, Roedl K, Mayer C, Brehm TT, van der Meirschen M, Hennigs A, Christopeit M, Fiedler W, Karagiannis P, Burdelski C, Schultze A, Huber S, Addo MM, Schmiedel S, Friese MA, Kluge S, and Schulze Zur Wiesch J. (2021) Multi-dimensional and longitudinal systems profiling reveals predictive pattern of severe COVID-19. iScience. 24:10275210.1016/j.isci.2021.102752.

Ahmadi P, Hartjen P, Kohsar M, Kummer S, Schmiedel S, Bockmann JH, Fathi A, Huber S, Haag F, and Schulze Zur Wiesch J. (2020) Defining the CD39/CD73 Axis in SARS-CoV-2 Infection: The CD73(-) Phenotype Identifies Polyfunctional Cytotoxic Lymphocytes. Cells. 9:10.3390/cells9081750.

Mitarbeiter:innen

Prof. Dr. Hans-Willi Mittrücker, Melina Birus, Peter Bradtke, Emily Valentina Madelaine Huth, Dr. Friederike Raczkowski, Jonas Rösch, Joanna Schmid, Dennis Yüzen

Forschungsprojekte

T-Zellen spielen eine zentrale Rolle in Immunantworten gegen Pathogene, sie sind aber auch entscheidend für die Entstehung von Autoimmunerkrankungen. In unserer Arbeitsgruppe verwenden wir verschiedene Mausmodelle für bakterielle Infektionen und für Autoimmunerkrankungen der Niere um die Funktion und Regulation von T-Zellen zu untersuchen.

Schwerpunkte sind:

● Die Funktion und Regulation von CD4 und CD8 T-Zellen in Infektionen mit bakterielen Pathogenen. Die Analysen finden in Infektionsmodellen für Listeria monocytogenes, Staphylococcus aureus und Salmonella typhimurium statt.

● Die Rolle des Transkriptionsfaktors Interferon Regulatory Factor 4 (IRF4) in der Aktivierung und Differenzierung von T-Zellen.

● Die Regulation der T-Zellantwort durch den IL-6 Rezeptor.

● Autoimmune Nierenerkrankungen und die Rolle von T-Zellen in der Entstehung und Aufrechterhaltung von Immunreaktionen gegen glomeruläre Strukturen.

● Die Bedeutung von durch Adenenin-Nukleotide aktivierbarer Ca2+ Kanäle in T-Zellen.

Forschungsnetzwerke

DFG: SFB 1328 “Adenine Nucleotides in Immunity and Inflammation” (A03: “Adenine nucleotide-modulated T cell differentiation and effector Functions”, Prof. H.-W. Mittrücker, Prof. Samuel Huber)

DFG: SFB 1192 „Immune-mediated glomerular diseases” (A4: “Autoreactive T cell responses in glomerulonephritis”)

DFG: KFO 296 „Feto-maternal immune cross talk: Consequences for maternal and offspring’s health” (P2 "Identification of viral determinants and maternal immune responses underlying influenza A virus disease severity during pregnancy“, Prof. G. Gabriel, Prof. H.-W. Mittrücker, Prof. P. Arck).

Ausgewählte Publikationen

Harberts A, Schmidt C, Schmid J, Reimers D, Koch-Nolte F, Mittrücker H-W*, Raczkowski F*. Interferon Regulatory Factor 4 controls effector functions of CD8+ memory T cells. (2021) Proc Natl Acad Sci USA. 118:e2014553118. (*shared last authorship)

Krebs CF, Reimers D, Zhao Y, Paust HJ, Bartsch P, Nuñez S, Rosemblatt MV, Hellmig M, Kilian C, Borchers A, Enk LUB, Zinke M, Becker M, Schmid J, Klinge S, Wong MN, Puelles VG, Schmidt C, Bertram T, Stumpf N, Hoxha E, Meyer-Schwesinger C, Lindenmeyer MT, Cohen CD, Rink M, Kurts C, Franzenburg S, Koch-Nolte F, Turner JE, Riedel JH, Huber S, Gagliani N, Huber TB, Wiech T, Rohde H, Bono MR, Bonn S, Panzer U*, Mittrücker H-W*. (2020) Pathogen-induced tissue-resident memory TH17 (TRM17) cells amplify autoimmune kidney disease. Sci Immunol. 5:eaba4163. (*shared last authorship)

Klinge S, Yan K, Reimers D, Brede KM, Schmid J, Paust HJ, Krebs CF, Panzer U, Hopfer H, Mittrücker H-W. (2019) Role of regulatory T cells in experimental autoimmune glomerulonephritis. Am J Physiol Renal Physiol. 316:F572-F581.

Mahnke J, Schumacher V, Ahrens S, Käding N, Feldhoff LM, Huber M, Rupp J, Raczkowski F, Mittrücker H-W. (2016) Interferon Regulatory Factor 4 controls TH1 cell effector function and metabolism. Sci Rep. 6:35521.

Hünemörder S, Treder J, Ahrens S, Schumacher V, Paust H-J, Menter T, Matthys P, Kamradt T, Meyer-Schwesinger C, Panzer U, Hopfer H, Mittrücker H-W. (2015) TH1 and TH17 cells promote crescent formation in experimental autoimmune glomerulonephritis. J Pathol. 237:62-71.

Raczkowski F, Ritter J, Heesch K, Schumacher V, Höcker L, Raifer H, Klein M, Bopp T, Harb H, Kesper DA, Pfefferle PI, Grusdat M, Lang PA, Mittrücker H-W*, Huber M*. (2013) The transcription factor Interferon Regulatory Factor 4 is required for the generation of protective effector CD8+ T cells. Proc Natl Acad Sci USA. 110:15019-15024. (*shared last authorship)

Mittrücker H-W, Steinhoff U, Köhler A, Krause M, Lazar D, Mex P, Miekley D, Kaufmann SHE. (2007) Poor correlation between BCG vaccination-induced T cell responses and protection against tuberculosis. Proc Natl Acad Sci USA. 104:12434-12439.

Kursar M, Höpken UE, Koch M, Köhler A, Lipp M, Kaufmann SHE, Mittrücker H-W. (2005) Differential requirements for the chemokine receptor CCR7 in T-cell activation during Listeria monocytogenes infection. J Exp Med. 201:1447-1457.

Kursar M, Bonhagen K, Fensterle J, Köhler A, Hurwitz R, Kamradt T, Kaufmann SHE, Mittrücker H-W. (2002) Regulatory CD4+ CD25+ T cells restrict memory CD8+ T cell responses. J Exp Med. 196:1585-1592.

Mittrücker H-W, Matsuyama T, Grossman A, Kündig TM, Potter J, Shahinian A, Wakeham A, Patterson B, Ohashi PS, Mak TW. (1997) Requirement for the transcription factor LSIRF/IRF4 for mature B and T lymphocyte function. Science. 275:540-543.

Kooperationspartner

Universitätsklinikum Hamburg-Eppendorf

Marcus Altfeld, Friedrich Koch-Nolte, Friedrich Haag, Eva Tolosa Institut für Immunologie Ulf Panzer, Christian Krebs, Oliver Steinmetz, Nicola Tomas, Elion Hoxha, Rolf Stahl III. Medizinische Klinik Samuel Huber, Nicola Gagliani, Peter Hübener I. Medizinische Klinik Petra Arck Geburtshilfe und Pränatalmedizin Gisa Tiegs Experimentelle Immunologie und Hepatologie Andreas Guse, Björn-Philipp Dierks Biochemie und Molekulare Zellbiologie Tim Magnus, Björn Rissiek Klinik für Neurologie Cathy Meyer-Schwesinger Institut für Physiologie Manuel Friese Institut für Neuroimmunologie und Multiple Sklerose

Externe Partner

Helmut Hopfer Universitätsspital Basel Minka Breloer, Hanna Lotter Bernhard-Nocht Institut für Tropenmedizin, Hamburg Magdalena Huber, Ulrich Steinhoff, Michael Lohoff Philipps-Universität Marburg Dirk Schmidt-Arras, Stefan Rose-John Christian-Albrechts Universität Kiel Santina Bruzzone University of Genova, Italy Gülsah Gabriel Leibniz Institute for Experimental Virology, Hamburg

Dissertationen

2021

Justus Mahnke (Dr. Med.) Der Transkriptionsfaktor Interferon Regulatory Factor 4 kontrolliert die Effektorfunktion und den Metabolismus von TH1-Zellen

Lucas Schumacher (Dr. Med.) Characterisation of single domain antibodies against the ADP-ribosylating Clostridium difficile toxin CDT and establishment of a murine infection model with the pathogen.

2019

Moritz Link (Dr. Med.) Die Bedeutung von ADAM17 für die TH1 und CD8 T-Zellantwort gegen bakterielle Krankheitserreger

Daniel Reimers (Dr. rer nat.) Der Einfluss von bakteriell induzierten geweberesidenten T-Zellen auf renale Autoimmunerkrankungen

2018

Alexandra Maximiliane Hierweger (Dr. rer. biol. hum. / PhD) Influence of influenza A virus infections on the maternal immune system during pregnancy

2017

Jule Gehrke (Master Infection Biology) The influence of TRPM2 on the CD8+ T‐cell response to Listeria monocytogenes

Karsten Yan (geb. Lücke) (Dr. rer nat.) Einfluss von IL‐6 auf die angeborene Immunantwort der Maus gegen Listeria monocytogenes

2016

Stefanie Klinge (geb. Ahrens) (Dr. rer. nat.) Die Rolle von regulatorischen T-Zellen in der autoimmunen anti-glomerulären Basalmembran-Glomerulonephritis

Kristina Broder (Dr. Med.) Die Regulation von CD8+ T-Zellen durch den Transkriptionsfaktor Interferon Regulatory Factor 4 (IRF4)

Mitarbeiter:innen

Prof. Friedrich Koch-Nolte, Dr. rer. nat. Anna Marei Mann, Dr. rer. nat. Julia Hambach, Marten Dannenberg, Anya Duttmann, Josephine Gebhardt, Saruul Jargalsaikhan, Luca Pape, Nathalie Richter, Henry Risch, Waldemar Schäfer, Alessa Schaffrath, Fabienne Seyfried, Natalie Tode

Forschungsprojekte

Die Arbeitsgruppe Molekulare Immunologie untersucht membranständige Rezeptoren und Enzyme von Immun- und Krebszellen und entwickelt Nanobodies und Antikörper gegen diese Zielstrukturen.

Forschungsnetzwerke

DFG: SFB 1192 „Immune-mediated glomerular diseases” (B05: Nanobody basierte Behandlungsstrategien bei Glomerulonephritiden, Nicola Wanner, Friedrich Nolte)

DFG: SFB 1328 “Adenine Nucleotides in Immunity and Inflammation” (A10: Rolle von Adenin-Nukleotiden und purinergen Rezeptoren im Fettgewebe, Jörg Heeren, Friedrich Nolte; Z02: Antikörper und Nanobodies für die Erforschung und Modulation des Adenin-Nukleotid-Signalings Friedrich Haag, Friedrich Nolte)

DFG: FOR 2879 "ImmunoStroke: From Immune Cells to Stroke Recovery" (A1: Die Rolle von ATP und NAD als Gefahrensignale und Therapieziel in der Erholungsphase des Schlaganfalles, Tim Magnus, Friedrich Nolte)

DFG/ANR: Evaluierung von P2X7 als therapeutisches Target in der Autoimmun-Enzephalitis und in der Tumorabwehr (Sahil Adriouch, Univ. Rouen, Friedrich Nolte)

DFG: Entwicklung von Nanobody-basierten bispezifischen Diabodies und Schwereketten-Antikörper für das gezielte Targeting von Myelomzellen (Peter Bannas, Radiologie, Friedrich Nolte)

BMBF COMMUTE Combinatorial and multidisciplinary targeting of effective gene therapy vectors (P5 Targeting glioblastoma and muscle dystrophy with Nanobody-displaying AAV, Friedrich Nolte)

Short CV Prof. Friedrich Koch-Nolte

Deputy Director (since 1997)

Phone: +49 (0) 40 7410 - 53612
Mail: nolte@uke.de

University Degrees
Biology, Wesleyan University 1976 (BA)
Medicine, University of Tübingen, 1983 (MD)
Molceular Biology, University of Hamburg, 1986 (Diploma)

Professor associée at the University of Rouen, France (2006-2007)
Visiting scientist at the University of California, San Francisco, CA, USA (1994)
Visiting scientist at the DNAX Research Institute of Molecular Biology, Palo Alto, CA, USA (1997)
Visiting scientist at The Jackson Lab, Bar Harbor, ME, USA (1991, 1999)

Forschungspreis der Hamburger Krebsgesellschaft (2019)

Mentorship Award of the Simon-Claussen Foundation (2009)
Research Award of the Werner Otto Foundation (1997)
Research Award of the Martini Foundation (1991)

Member of the
Deutsche Gesellschaft für Immunologie (DGfI)
Deutsche Gesellschaft für Biochemie und Molekularbiologie (GBM)
Deutsche Gesellschaft für Zellbiologie (DGZ)

Kooperationspartner

Universitätsklinikum Hamburg-Eppendorf

Friedrich Haag, Eva Tolosa, Hans-Willi Mittrücker Institut für Immunologie Andreas Guse, Jörg Heeren, Ralf Fliegert Biochemie und Molekulare Zellbiologie Peter Bannas Klinik für Radiologie Tim Magnus, Björn Rissiek Klinik für Neurologie Cathy Meyer-Schwesinger Institut für Physiologie Nicola Wanner, Nicola Tomas, Elion Hoxha, Rolf Stahl, Ulf Panzer III. Medizinische Klinik Boris Fehse, Kristoffer Rieken Experimentelle Immunologie und Hepatologie Irm Hermanns Borgmeyer Zentrum für Molekulare Neurobiologie Katja Weisel, Walter Fiedler I. Medizinische Klinik Thomas Braulke Kinderbiochemie

Externe Partner

Sahil Adriouch Universität Rouen Michael Hottiger Universität Zürich Pablo Pelegrin Universiät Murcia Mascha Binder Universiät Halle Martin Trepel Universiät Augsburg Dirk Grimm Universiät Heidelberg Annette Nicke Ludwig-Maximilians-Universität München Carsten Watzl Universiät Dortmund Florian Schmidt Universität Bonn Francesco Di Virgilio, Elena Adinolfi Universiät Ferrara Santina Bruzzone, Paolo Malatesta Universiät Genua Jan van Lier Universiät Amsterdam Jaime Sancho Universiät Granada Pablo Engel Universiät Barcelona Ursula Dietrich Georg-Speyer-Haus William Heath Universiät Melbourne William Petri Universiät Virginia Steven Mansoor Universiät Oregon Hidde Ploegh Boston Children's Hospital

Research Interests

The focus of our laboratory is on the molecular characterization of lymphocyte membrane proteins, in particular receptors and enzymes involved in signaling by extracellular nucleotides. We generate monoclonal and single domain antibodies as new research and therapeutic tools. We are interested in ADP-ribosylation as a pathogenic mechanism of bacterial toxins and as a reversible posttranslational modification regulating protein functions. Using genetic immunization and antibody engineering, we strive to develop new tools for combating infections and for treating diseases of the immune system.

1. Lymphocyte membrane proteins: enzymes and receptors involved in signaling by extracellular NAD and ATP

Membrane proteins mediate the communication of cells with their environment. They function as receptors for soluble ligands and counter-receptors on other cells, as ion channels, nutrient transporters, and enzymes. The nucleotides NAD and ATP are key metabolites of energy metabolism found in cells from all kingdoms of life. The cell membrane is impermeable to these nucleotides, but they can exit cells via channels or pores gated by mechanical and/or electrical stimuli. During infection and inflammation injured cells release NAD and ATP through the damaged cell membrane. Extracellular NAD and ATP alert cells of the immune system to sites of tissue damage.

Cells of the immune system are equipped with a variety of sensors for these nucleotides (Fig. 1), including ligand-gated ion channels (P2X purine receptors) and nucleotide-metabolizing ecto-enzymes (CD38, CD296). CD296 (ARTs) functions as NAD-sensors and relay information about the levels of extracellular NAD into ADP-ribosylation of cell surface and secreted proteins. The NAD-hydrolyzing ecto-enzyme CD38 restricts the intensity and duration of NAD-signaling in the extracellular space by hydrolyzing NAD. Opening of the P2X7 ion channel is induced by binding of the soluble ligand ATP, or by NAD-dependent ADP-ribosylation. Passage of ions through P2X7 (calcium into the cell, potassium out of the cell) triggers a cascade of downstream events, including protease activation (caspases and ADAMs), externalization of phosphatidylserine (PS), activation of the inflammasome and cell death. Mice that cannot express the mentioned purine receptors or ecto-enzymes show impaired immune responses. The receptors and enzymes of purinergic signaling, thus, present potential targets for new anti-inflammatory or immune-stimulating drugs. In mouse models, activating the ARTC2 > P2X7 axis can enhance anti-tumor responses, while blocking this axis can reduce inflammation in autoimmune diseases.

2. Antibody engineering: genetic immunization, single domain antibodies

Antibodies are important tools of research, diagnosis and therapy. Antibodies are raised by immunizing experimental animals with proteins, synthetic peptides, or DNA. In the case of DNA immunizations, the protein encoded by the DNA is produced in its native conformation by the cells of the immunized animal and the induced immune response yields antibodies directed against proteins in native conformation (ADAPINCs) (Fig. 2). Such antibodies are useful in many applications were anti-peptide antibodies fail, e.g. affinity purification, flow cytometry, ELISA, FACS, and functional studies. cDNA immunization is offered as a service via the antibody core facility of the UKE.

Llamas produce unusual antibodies composed only of heavy chains. Their antigen-binding domain (VHH) is readily produced as a soluble recombinant protein, also designated nanobody or single domain antibody (sdAb) (Fig. 3). Nanobodies can form finger like extrusions that block clefts on protein surfaces, such as the active site crevice of enzymes, the ligand binding domain of a receptor, and the receptor binding domain of a virus. Nanobodies have great potential as therapeutics and as imaging agents.

We have generated enzyme-blocking nanobodies from llamas immunized with different ARTs: the SpvB Salmonella toxin, the binary clostridium difficile toxin CDTa, and the T cell ecto-enzyme ARTC2 (CD296). These nanobodies protect cells from the cytotoxic effects of SpvB, CDT, and ARTC2. In case of ARTC2, the nanobodies effectively block ARTC2 on the cell surface of regulatory T cells and iNKT cells within 10 minutes after intravenous injection. These nanobodies provide an important tool for protecting these regulatory cells from death induced by NAD released during tissue preparation.
We have also generated nanobodies against CD38, the major NAD-hydrolyzing ecto-enzyme. CD38 is emerging as a therapeutic target in multiple myeloma and other hematopoertic malignancies. Some of our nanobodies outperform the recently licensed CD38-specific monoclonal antibody Daratumumab (Darzalex) in cytotoxicity vs. hematopoetic cancer cell lines.
In cooperation with Ablynx, a Belgian company devoted to developing nanobodies for human therapies, we have generated nanobodies that block or enhance gating of the P2X7 ion channel. The P2X7-blocking nanobodies ameliorated inflammation in mouse models of glomerulonephritis and allergic dermatitis. In endotoxin-treated human blood cells, they effectively blocked LPS/ATP-induced release of the potent pro-inflammatory cytokine IL-1ß.
In order to facilitate the generation of new nanobodies, we have cloned the nanobody-encoding IgH locus from llama and successfully transferred an engineered version of this locus to transgenic mice (Fig. 4). Upon immunization, these mice produce nanobody-based heavy chain antibodies that undergo somatic hypermutation and class switch recombination. This novel platform greatly expands our capacity to generate functional nanobodies against interesting targets.

3. Posttranslational modifications: ADP-ribosylation, glycosylation, lipid anchors

The function of proteins can be regulated via the attachment of chemical moieties. These enzyme-catalyzed modifications, coined posttranslational modifications (PTMs), include phosphorylation, glycosylation, ADP-ribosylation, and the attachment of lipid anchors.
ADP-ribosylation is a reversible PTM, in which ADP-ribosyltransferases (ARTs) transfer the ADP-ribose moiety from NAD onto a specific amino acid side chain in a target protein and ADP-ribosylhydrolases (ARHs) remove the ADP-ribose group (Fig. 5). We have determined the 3D structures of a prototype ART and a prototype ARH. The 3D structure of rat ARTC2 resembles a pacman with a wide-open mouth crunching on the substrate NAD. The 3D structure of human ARH3 resembles a pumpkin in which four alpha helices coordinate two magnesium ions at the bottom of the active site crevice.

ADP-ribosylation is used by pathogenic toxins such as Diphtheria, Pertusssis and Clostridial toxins to modulate host protein functions. Toxin-related ARTs are expressed by cells of the immune system. ADP-ribosylation of membrane proteins can be monitored using labeled analogues of NAD. Using 32P-NAD, ADP-ribosylation results in radiolabeling of the target protein. Using etheno-NAD, ADP-ribosylation of target proteins can be detected with a monoclonal antibody directed against etheno-adenosine. This 1G4 antibody can be used to sort cells on the basis of cell surface ART-activity and to purify etheno-ADP-ribosylated proteins.

ARTC2 itself is posttranslationally modified, e.g. by attachment of a GPI lipid anchor (Fig. 1 above). The GPI-anchor targets ARTC2 to lipid rafts, specialized regions of the cell membrane that play an important role in signal transduction, e.g. during activation of T cells by antigen presenting cells. The association of ARTC2 with lipid rafts focuses ARTC2 onto specific target proteins and thereby may regulate the signaling function of raft-associated proteins. ADP-ribosylation of the P2X7 ion channel on arginine residue 125 activates P2X7 to form a non-selective ion-channel, permitting calcium ions to enter the cell and potassium ions to exit the cell. This induces dramatic alterations of the cell membrane, including the externalization of PS, shedding of L-selectin, and formation of membrane blebs.

Ausgewählte Publikationen

reviews

Eggers M, Rühl F, Haag F, Koch-Nolte F. (2021) Nanobodies as probes to investigate purinergic signaling. Biochem Pharmacol. 87:114394. review PMID: 33388283.

Linden J, Koch-Nolte F, Dahl G. (2019) Purine Release, Metabolism, and Signaling in the Inflammatory Response. Annu Rev Immunol. 37:325-347. review. PMID: 30676821

Stortelers C, Pinto-Espinoza C, Van Hoorick D, Koch-Nolte F. (2018) Modulating ion channel function with antibodies and nanobodies. Curr Opin Immunol. 52:18-26. Review PMID: 29579624

Bannas P, Hambach J, Koch-Nolte F. (2017) Nanobodies and Nanobody-Based Human Heavy Chain Antibodies As Antitumor Therapeutics. Front Immunol. 8:1603. Review PMID: 29213270

Wesolowski J, Alzogaray V, Reyelt J, Unger M, Juarez K, Urrutia M, Cauerhff A, Danquah W, Rissiek B, Scheuplein F, Schwarz N, Adriouch S, Boyer O, Seman M, Licea A, Serreze DV, Goldbaum FA, Haag F, Koch-Nolte F. (2009) Single domain antibodies: promising experimental and therapeutic tools in infection and immunity. Med Microbiol Immunol. 198:157-74. Review PMID: 19529959

Koch-Nolte F, Kernstock S, Mueller-Dieckmann C, Weiss MS, Haag F. (2008) Mammalian ADP-ribosyltransferases and ADP-ribosylhydrolases. Front Biosci. 13:6716-29. PMID: 18508690

research reports

Hambach J, Stähler T, Eden T, Wendt D, Tode N, Haag F, Tolosa E, Altfeld M, Fathi A, Dahlke C, Addo MM, Menzel S, Koch-Nolte F. (2021) A simple, sensitive, and low-cost FACS assay for detecting antibodies against the native SARS-CoV-2 spike protein. Immun Inflamm Dis. 9:905-917. PMID: 33979020.

Schriewer L, Schütze K, Petry K, Hambach J, Fumey W, Koenigsdorf J, Baum N, Menzel S, Rissiek B, Riecken K, Fehse B, Röckendorf JL, Schmid J, Albrecht B, Pinnschmidt H, Ayuk F, Kröger N, Binder M, Schuch G, Hansen T, Haag F, Adam G, Koch-Nolte F, Bannas P. (2020) Nanobody-based CD38-specific heavy chain antibodies induce killing of multiple myeloma and other hematological malignancies. Theranostics. 10:2645-2658. PMID: 32194826

Eichhoff AM, Börner K, Albrecht B, Schäfer W, Baum N, Haag F, Körbelin J, Trepel M, Braren I, Grimm D, Adriouch A, Koch-Nolte F. (2019) Nanobody-enhanced targeting of AAV gene therapy vectors Mol Ther Methods Clin Dev. 15:211-220. PMID: 31687421

Eden T, Menzel S, Wesolowski J, Bergmann P, Nissen M, Dubberke G, Seyfried F, Albrecht B, Haag F, Koch-Nolte F. (2018) A cDNA Immunization Strategy to Generate Nanobodies against Membrane Proteins in Native Conformation. Front Immunol. 8:1989. PMID: 29410663

Danquah W, Meyer-Schwesinger C, Rissiek B, Pinto C, Serracant-Prat A, Amadi M, Iacenda D, Knop JH, Hammel A, Bergmann P, Schwarz N, Assunção J, Rotthier W, Haag F, Tolosa E, Bannas P, Boué-Grabot E, Magnus T, Laeremans T, Stortelers C, Koch-Nolte F. (2016) Nanobodies that block gating of the P2X7 ion channel ameliorate inflammation. Sci Transl Med. 8:366ra162. PMID: 27881823

Adriouch S, Bannas P, Schwarz N, Fliegert R, Guse AH, Seman M, Haag F, Koch-Nolte F. (2008) ADP-ribosylation at R125 gates the P2X7 ion channel by presenting a covalent ligand to its nucleotide binding site. FASEB J. 22:861-9. PMID: 17928361

Koch-Nolte F, Reyelt R, Schössow B, Schwarz N, Scheuplein F, Rothenburg S, Haag F, Alzogaray V, Cauerhff A, Goldbaum F. (2007) Single domain antibodies from llama effectively and specifically block T cell ecto-ADP-ribosyltransferase ART2.2 in vivo. FASEB J. 21:3490-8. PMID: 17575259

Mueller-Dieckmann C, Kernstock S, Lisurek M, von Kries JP, Haag F, Weiss MS, Koch-Nolte F. (2006) The structure of human ADP-ribosylhydrolase 3 (ARH3) provides insights into the reversibility of protein ADP-ribosylation. Proc Natl Acad Sci U S A. 103:15026-31. PMID: 17015823

Otto H, Reche PA, Bazan F, Dittmar K, Haag F, Koch-Nolte F. (2005) In silico characterization of the family of PARP-like poly(ADP-ribosyl)transferases (pARTs). BMC Genomics. 6:139. PMID: 16202152

Bannas P, Adriouch S, Kahl S, Braasch F, Haag F, Koch-Nolte F. (2005) Activity and specifity of toxin-related mouse T cell ecto-ADP-ribosyltransferase ART2.2 depends on it association with lipid rafts. Blood. 105:3663-70. PMID: 15657180

Seman M, Adriouch S, Scheuplein F, Krebs C, Freese D, Glowacki G, Deterre P, Haag F, Koch-Nolte F. (2003) NAD-induced T cell death: ADP-ribosylation of cell surface proteins by ART2 activates the cytolytic P2X7 purinoceptor. Immunity. 19:571-82. PMID: 14563321

Otto H, Tezcan-Merdol D, Girisch R, Haag F, Rhen M, Koch-Nolte F. (2000) The spvB gene-product of the Salmonella enterica virulence plasmid is a mono(ADP-ribosyl)transferase. Mol Microbiol. 37:1106-15. PMID: 10972829

Koch-Nolte F, Petersen D, Balasubramanian S, Haag F, Kahlke D, Willer T, Kastelein R, Bazan F, Thiele HG. (1996) Mouse T cell membrane proteins Rt6-1 and Rt6-2 are arginine/protein mono(ADPribosyl)transferases and share secondary structure motifs with ADP-ribosylating bacterial toxins. J Biol Chem. 271:7686-93. PMID: 8631807

Koch F, Haag F, Kashan A, Thiele HG. (1990) Primary structure of rat RT6.2, a nonglycosylated phosphatidylinositol-linked surface marker of postthymic T cells. Proc Natl Acad Sci U S A. 87:964-7. PMID: 2300588

Koch F, Thiele HG, Low MG. (1986) Release of the rat T cell alloantigen RT-6.2 from cell membranes by phosphatidylinositol-specific phospholipase C. J Exp Med. 164:1338-43. PMID: 3489808

Mitarbeiter:innen

Prof. Felix Stahl, Lama Al Jurdi, Luis Fonseca Brito, Dorottya Schöffer, Silvia Tödter

Forschungsprojekte

Forschungsnetzwerke

DFG: KFO 296 „Feto-maternal immune cross talk: Consequences for maternal and offspring’s health”

Ausgewählte Publikationen

Fonseca Brito L, Ostermann E, Perez A, Tödter S, Virdi S, Indenbirken D, Glau L, Gieras A, Brixel R, Arens R, Grundhoff A, Arck P, Diemert A, Tolosa E, Brune W, Stahl FR. (2024) Limited protection against early-life cytomegalovirus infection results from deficiency of cytotoxic CD8 T cells. bioRxiv. 2024.07.10.602923.

Fonseca Brito L, Todter S, Kottlau J, Cermann K, Spier A, Petersen E, Schafer I, Twerenbold R, Aepfelbacher M, Lutgehetmann M, Stahl FR. Performance of an interferon-gamma release assay-based test for cell-mediated immunity to SARS-CoV-2. (2023) Front Immunol. 14:1069968.

Fonseca Brito L, Brune W, Stahl FR. Cytomegalovirus (CMV) Pneumonitis: Cell Tropism, Inflammation, and Immunity. (2019) Int J Mol Sci. 20(16).

Stahl FR, Jung R, Jazbutyte V, Ostermann E, Todter S, Brixel R, Kemmer A, Halle S, Rose-John S, Messerle M, Arck PC, Brune W, Renne T. (2018) Laboratory diagnostics of murine blood for detection of mouse cytomegalovirus (MCMV)-induced hepatitis. Sci Rep. 8(1):14823.

Lueder Y, Heller K, Ritter C, Keyser KA, Wagner K, Liu X, Messerle M, Stahl FR#, Halle S#, Forster R#. Control of primary mouse cytomegalovirus infection in lung nodular inflammatory foci by cooperation of interferon-gamma expressing CD4 and CD8 T cells. (2018) PLoS Pathog. 14(8):e1007252.

Halle S, Keyser KA, Stahl FR, Busche A, Marquardt A, Zheng X, Galla M, Heissmeyer V, Heller K, Boelter J, Wagner K, Bischoff Y, Martens R, Braun A, Werth K, Uvarovskii A, Kempf H, Meyer-Hermann M, Arens R, Kremer M, Sutter G, Messerle M, Forster R. (2016) In Vivo Killing Capacity of Cytotoxic T Cells Is Limited and Involves Dynamic Interactions and T Cell Cooperativity. Immunity. 44(2):233-45.

Stahl FR#, Keyser KA#, Heller K, Bischoff Y, Halle S, Wagner K, Messerle M, Forster R. Mck2-dependent infection of alveolar macrophages promotes replication of MCMV in nodular inflammatory foci of the neonatal lung. (2015) Mucosal Immunol. 8(1):57-67.

Stahl FR, Heller K, Halle S, Keyser KA, Busche A, Marquardt A, Wagner K, Boelter J, Bischoff Y, Kremmer E, Arens R, Messerle M, Forster R. (2013) Nodular inflammatory foci are sites of T cell priming and control of murine cytomegalovirus infection in the neonatal lung. PLoS Pathog.9(12):e1003828.

#shared authorships