1) Name – Title of the research project in CCHD
Hannes Stockinger – Early signalling events in T cell activation
2) Coordinates of the Faculty Member
Centre of Physiology, Pathophysiology and Immunology, Medical Univ. Vienna, Lazarettgasse 19, A-1090 Vienna; Tel +43 1 40160 33001; Fax +43 1 40160 933002;
eMail: hannes.stockinger@meduniwien.ac.at.
3) Keywords
adaptive immunity, T cell activation, signal transduction, protein kinase, imaging
4) Research interest of the Faculty Member
Dr. Stockinger is interested in the structure and function of surface receptors on T cells and accessory/dendritic cells to identify novel targets for influencing abnormal and unwanted immune reactions in immunological disorders and diseases. Internationally well recognized are the investigations, which contribute to the understanding of how glycosylphosphatidylinositol (GPI)-anchored receptor proteins transduce signals across the plasma membrane. These studies were fundamental for the identification and characterization of special membrane microdomains, called lipid rafts, which are more and more believed to control initiation of signal transduction across the membrane of cells. Currently, Dr. Stockinger puts special emphasis is on the development of novel microscopic techniques to analyze the dynamic of receptors and signalling molecules on the single molecule level in living cells in real time. First results derived from this “real-time biochemistry with single molecules in living cells” suggest a new vista of how molecules function in cells. Based on these studies new diagnostic assays will be developed.
5) Collaborations within CCHD
With B.R. Binder, there is a long lasting collaboration in respect of signal transduction of the urokinase plasminogen activator receptor CD87 and regulation of its function in fibrinolysis, cell adhesion and cell migration (7 joint publications). Currently, the main joint project with B.R. Binder in close collaboration with D. Kerjaschki is focused on elucidating the function of the lymphatic endothelial marker podoplanin in vascular biology and immunity (1 manuscript submitted, 1 manuscript in revision). With S. Böhm, live FRET experiments, with T. Stulnig long-lasting collaboration on integrity, composition and function of lipid rafts (5 joint publications).
6) Collaborating research groups where PhD Students can perform their research stay
Dr. Giuseppina Caligiuri, INSERM UMRS 681, Université Pierre et Marie Curie Paris 6, 15, rue de l'Ecole de Mèdecine, F-75006 Paris, France
Prof. Dr. Vaclav Horejsi, Director, Institute of Molecular Genetics, Academy of Sciences of the Czech Republic, Videnska 1083, 142 20 Prague 4, Czech Republic
Prof. Dr. Watchara Kasinrerk, Clinical Immunology Branch, Department of Medical Technology, Faculty of Associated Medical Sciences, Chiang Mai University, Chiang Mai 50200, Thailand.
Prof. Dr. Gerhard Schütz, Biophysics Institute, Johannes Kepler University Linz, Altenbergerstr. 69, A-4040 Linz, Austria
Prof. Dr. Kai Simons, Max Planck Institute of Molecular Cell Biology and Genetics, Pfotenhauerstr. 108, D-01307 Dresden, Germany
Prof. Dr. Michael Stürzl, Division of Molecular and Experimental Surgery, Department of Surgery, University of Erlangen-Nuremberg, Schwabachanlage 10, 91054 Erlangen, Germany.
Dr. Ulrich H. Weidle, Drug Development Oncology, Roche Diagnostics GmbH, Pharma Research, Nonnenwald 2, 82377 Penzberg, Germany (www.roche.de).
7) Industrial partners (if applicable)
Univ. Doz. Dr. Birgit Reipert, Director Immunology, Baxter Austria (www.baxter.at)
8) Know-how and infrastructure of the research group
The Department of Dr. Stockinger has outstanding expertise in immune cell biology with major interest towards negative regulating receptors and signalling pathways in T cells to correct pathological and unwanted immune reactions occurring in autoimmunity, allergy, transplantation. Recent advances are: In vivo confirmation of glycoprotein CD31 as negative regulator of T cells by curing atherosclerosis in mice (Groyer et al., 2007); regulation of integrin LFA-1- and CD43-mediated adhesion of T cells and other leukocytes by glycoprotein CD147 (Khunkaewla et al., 2008); identification of a molecular pathway how the negative regulating GBP-1 inhibits cell function (Weinländer et al., 2008); the mannose-6-phosphate insulin-like growth factor II receptor (CD222) as negative regulator of fibrinolysis and cell migration (Prager et al., 2004; Leksa et al, 2005, Schiller et al., 2008, Probst et al., 2009), large scale production of cytotoxic T cells for adoptive immunotherapy (Jursik et al., 2009). As tools for these experiments, we generated a large panel of monoclonal antibodies to structurally and functionally characterize the target molecules by molecular biological, cellular and biochemical assays. For the latter approaches we established a number of “gain of function” and “loss of function” assays by using recombinant and tagged forms as well as siRNA sequences of the target molecules. Furthermore, a number of immunobiochemical assays including affinity chromatography, quantitative and qualitative immunoprecipitation, Western blotting are established.
A recent research topic of the Department is the development of advanced microscopy techniques to analyze the function of signalling molecules directly in living cells. For this purpose we mastered construction of fluorescent protein (FP) tagged receptors and signalling molecules. For instance, we have available a number of FP variants of Lck [the major Src protein tyrosine kinase (SFK) of T cells], G-protein GBP-1 and transmembrane receptors. Because FPs are with 27 kilodalton relatively large, special attention has to be given to keep the functionality of the tagged molecules. We know now both vulnerable and susceptible sites to modulate and tag Lck and other members of the SFK family. This knowledge is an invaluable cache to tag any other kind of molecule with FPs. These biological tools were the prerequisite for the ultra-sensitive microscopy techniques that we developed in parallel to analyze the function of signalling molecules directly in living T cells (Moertelmaier et al., 2005; Madl et al., 2006, Drbal et al., 2007; Wieser et al., 2007a; Wieser et al., 2007b). The most successful development is 1) a FRET-based biosensor of Lck that can report changes in the functional state of the protein in living cells (Paar et al., 2008; Paster et al., 2009), 2) single molecule tracking in living cells to get insight into size and function of receptor complexes (Drbal et al., 2007; Wieser et al., 2007a), and 3) live cell micropatterning that allows to screen for and characterize molecular interactions directly in the living cell (Schwarzenbacher et al., 2008).
The results were published in approximately 140 scientific papers, a variety of them in the very top journals such as Cell, Science, Nature Methods and The Journal of Experimental Medicine that were cited nearly 6000 times. Five patents are currently active.
The Department is also deeply involved in the education programs of the MUV. Besides giving a number of lectures and seminars for undergraduate students, the staff of the Department is well prepared in training of graduate students and in post-graduate education. Further, the chairman of the Department is the dean of the PhD-Programmes at the MUV.
