Recreating biological phenomena in model systems with defined components and properties has proven to be a powerful means for dissecting molecular mechanisms. Our research is dedicated to the development of such model systems that allow the reconstitution, observation and physical manipulation of cellular processes down to the single molecule level. In particular, we apply micro- and nanostructured biointerfaces to gain a quantitative and mechanistic understanding of plasma membrane organization and associated cellular signaling processes, with a particular interest in the molecular mechanisms of T-cell signaling.
DNA origami as a tool to study cell surface receptor signaling
he nanoscale spatial organization of ligands and receptors is emerging as an important theme in regulating cell behavior. We have developed a cell-responsive biomimetic interface based on laterally mobile functionalized DNA origami platforms to probe the spatial requirements for receptor-mediated signaling. While fully responsive to dynamic receptor rearrangements on the cell surface upon ligand engagement, the biointerface allows the experimenter to adjust protein distances with nanoscale precision as a means to disturb signaling. We currently apply and refine our biointerface to identify and validate cell biological, biophysical and biochemical parameters underlying sensitized T-cell antigen detection.
Ligand-functionalized DNA origami platforms are attached via cholesterol-modified oligonucleotides to supported lipid bilayers, where they diffuse freely. The platforms set a minimum distance δ between neighboring ligands (e.g. pMHCs)as they cluster after T-cell receptor (TCR) engagement.
Protein micropatterning to study plasma membrane organization and function
It is inherently difficult to study the interactions of proteins and lipids in the plasma membrane of living cells. We have developed a protein micropatterning assay where we use microstructured surfaces to confine a protein of interest to specific areas in the cellular plasma membrane and monitor the interaction with other membrane constituents via single molecule microscopy. By doing so we can extract kinetic parameters of specific interactions but also study fundamental elements of plasma membrane organization.
a, Microstructuring of proteins in the live cell plasma membrane. b, Mammalian cell on a micropatterned surface; the patterned protein is labeled with mGFP. c, Monitoring the interaction of lipids with micropatterned proteins via single molecule tracking of lipids.
R. Platzer, J. Hellmeier, J. Göhring, I. Doel Perez, P. Schatzlmaier, C. Bodner, M. Focke-Tejkl, G.J. Schütz, E. Sevcsik, H. Stockinger, M. Brameshuber, J.B. Huppa (2023). Monomeric agonist peptide/MHCII complexes activate T-cells in an autonomous fashion. EMBO Reports e57842
J. Hellmeier, R. Platzer, J.B. Huppa, E. Sevcsik* (2023). A DNA origami-based biointerface to interrogate the spatial requirements for sensitized T-cell antigen recognition. Methods Mol. Biol. 2654, 277
I. Hartl, V. Brumovska, Y. Strieder, A. Yasari, G.J. Schütz, E. Sevcsik*, I. Tiemann-Böge* (2023) Measurement of FGFR3 signaling at the cell membrane via total internal reflection fluorescence microscopy to compare the activation of FGFR3 mutants. J. Biol. Chem. 299, 102832
J. Hellmeier, R. Platzer, V. Mühlgrabner, M.C. Schneider, E. Kurz, G.J. Schütz, J.B. Huppa, E. Sevcsik* (2021). Strategies for the site-specific decoration of DNA origami nanostructures with functionally intact proteins. ACS Nano 15, 15057 doi: 10.1021/acsnano.1c05411
A. Schromm, L. Paulowski, Y. Kaconis, F. Kopp, M. Koistinen, A. Donoghue, S. Keese, C. Nehls, J. Wernecke, P. Garidel, E. Sevcsik, K. Lohner, S. Sanchez-Gomez, G. Martinez-de-Tejada, K. Brandenburg, M. Brameshuber, G.J. Schütz, J. Andrä, T. Gutsmann (2021). Cathelicidin and PMB neutralize endotoxins by multifactorial mechanisms including LPS interaction and targeting of host cell membranes. PNAS 118, e2101721118
J. Hellmeier, R. Platzer, A.S. Eklund, T. Schlichthärle, A. Karner, M.C. Schneider, V. Motsch, E. Kurz, V. Bamieh, M. Brameshuber, J. Preiner, R. Jungmann, H. Stockinger, G.J. Schütz, J.B. Huppa, E. Sevcsik* (2021). DNA origami demonstrate the unique stimulatory power of single pMHCs as T-cell antigens. PNAS 118, e2016857118
R. Platzer, B.K. Rossboth, M.C. Schneider, E. Sevcsik, F. Baumgart, H. Stockinger, G.J. Schütz, J.B. Huppa, M. Brameshuber (2020). Unscrambling fluorophore blinking for comprehensive cluster detection via photoactivated localization microscopy. Nature Commun. 11, 4993
A. M. Arnold, A.W.A.F. Reismann, E. Sevcsik, G.J. Schütz (2020). Monte Carlo simulations of protein micropatterning in biomembranes: effects of immobile nanofeatures with reduced diffusivity. J. Phys. D: Appl. Phys. 53, 435401
T. Váradi, M. Schneider, E. Sevcsik, D. Kiesenhofer, F. Baumgart, G. Batta, T. Kovács, R. Platzer, J.B. Huppa, J. Szöllősi, G.J. Schütz, M. Brameshuber, P. Nagy (2019). Homo- and heteroassociations drive activation of ErbB3. Biophys. J. 117, 1935
V. Motsch, M. Brameshuber, F. Baumgart, G.J. Schütz, E. Sevcsik* (2019). A micropatterning platform for quantifying interaction kinetics between the T cell receptor and an intracellular binding protein. Scientific Reports 9, 3288
M. Lindner, A. Tresztenyak, G. Fülöp, W. Jahr, A. Prinz, I. Prinz, J. Danzl, G.J. Schütz, E. Sevcsik* (2019). A fast and simple contact printing approach to generate 2D protein nanopatterns. Frontiers in Chemistry 6, 655
R. Hager, A.M. Arnold, E. Sevcsik, G.J. Schütz and S. Howorka (2018). Tunable DNA hybridization enables spatially and temporally controlled surface-anchoring of biomolecular cargo. Langmuir 34, 15021-15027
G. Fülop, M. Brameshuber, A.M. Arnold, G.J. Schütz, E. Sevcsik* (2018). Determination of the membrane environment of CD59 in living cells. Biomolecules 8, 28
M. Brameshuber, F. Kellner, B.K. Rossboth, H. Ta, K. Alge, E. Sevcsik, J. Göhring, M. Axmann, F. Baumgart, N.R.J. Gascoigne, S.J. Davis, H. Stockinger, G.J. Schütz and J.B. Huppa (2018). Monomeric TCRs drive T cell antigen recognition. Nature Immun. 19, 487-496
G.J. Schütz, J. Weghuber, P. Lanzerstorfer, E. Sevcsik* (2017). Protein Micropatterning Assay: Quantitative Analysis of Protein–Protein Interactions. Methods Mol. Biol. 1550, 261-270
A.M. Arnold, E. Sevcsik, G.J. Schütz (2016). Monte Carlo simulations of protein micropatterning in biomembranes: effects of immobile sticky obstacles. J. Phys. D: Appl. Phys. 49, 364002
E. Sevcsik* and G.J. Schütz* (2016). With or without rafts? Alternative views on cell membranes. Bioessays 38, 129-139
M. Brameshuber, E. Sevcsik, B. K. Rossboth, C. Manner, HP. Deigner, B. Peksel, M. Péter, Z. Török, A. Hermetter, G. J. Schütz (2016). Oxidized phospholipids inhibit the formation of cholesterol-dependent plasma membrane nanoplatforms. Biophys. J. 110, 205-213
E. Sevcsik*, M. Brameshuber, M. Fölser, J. Weghuber, A. Honigmann, G.J. Schütz (2015). GPI-anchored proteins do not reside in ordered domains in the live cell plasma membrane. Nature Commun. 6, 6969
Assistant Prof. Dipl.-Ing.in Dr.in techn. Eva Sevcsik
Head of Micro- and Nanostructured Biointerfaces Group, Research Unit of Biophysics