Ao.Univ.Prof. DI Dr. Johannes A. Schmid
Medical University of Vienna
Physiology Building, 3rd floor
Schwarzspanierstr. 17, 1090 Vienna, Austria
Know-how and research interests
Current research interests focus on inflammation, cancer and endothelial cell biology. Signal transduction pathways of inflammation are studied with a special focus on the NF-kappa B signaling, as well as interconnections with other signaling processes and transcription factors. In this context, my group is interested in cooperativity of effector molecules and signaling pathways representing an important aspect in multi-factorial diseases – and moreover providing a molecular basis for potential drug combination strategies.
Signal transduction networks are studied with a variety of experimental systems, such as cell culture of primary and transformed cells or transgene mouse models. Transfection and viral transduction methods are applied to achieve either ectopic expression or gene suppression of effector molecules followed by analyzing a variety of biological readouts such as cell proliferation, apoptosis or activation of cells or specific pathways. Moreover, we investigate the dynamics of signaling molecules (for instance nucleocytoplasmic shuttling) in living cells using fluorescent protein chimeras and various high-end laser scanning microscopy techniques (such as fluorescence loss in photobleaching, FLIP, or fluorescence recovery after photobleaching, FRAP). Interactions between proteins are studied by fluorescence resonance energy transfer (FRET) microscopy and verified with classical biochemical methods such as co-immunoprecipitation. Furthermore, we investigate protein-DNA interactions using classical methods (e.g. EMSA) as well as more sophisticated techniques such as chromatin immunoprecipitation (ChIP), avidin-biotin-complex-DNA assays (ABCD assays) or FRET. In addition to cell culture systems, we establish and exploit transgene mouse models using cell type specific expression or knock-out of genes of interest using the Cre-loxP technique and we study cooperativity of genes by cross-breeding of genetically modified mouse strains. Besides analyses of experimental model systems, we also investigate patient tissue samples using tissue arrays and novel methods of quantitative tissue cytometry to study correlations between candidate effector molecules.
Molecular signal transduction of the NF-kappa B pathway and its connections and cooperativity with other signaling pathways
Techniques and infrastructure of the research group
- classical methods of biochemistry and molecular biology
- microbiological techniques with bacteria and yeast
- different cloning strategies
- cultivation of human cells
- Transgene mouse models
- transient and stable transfections of human cells
- lentiviral gene transduction and shRNA mediated knock-down
- yeast two-hybrid and yeast one-hybrid screening
- immunoprecipitation and co-immunoprecipitation assays
- immunofluorescence microscopy
- confocal laser scanning microscopy
- subcellular fractionations by ultracentrifugation
- cryosectioning and paraffin sectioning of different tissues
- fluorometry (including wavelength and timescans)
- studies on fluorescence resonance energy transfer (FRET)
- flow analysis and cell sorting
- chromatography (size exclusion and others)
- kinase assays
- reporter gene assays
- apoptosis assays (in situ nick translation, TUNEL assay, PI staining and FACS)
- cell cycle analyses
- realtime PCR (ABI StepOne Plus, 96-well)
- Western Blot Imager
- Fluorescence microscopes
- Laser Scanning microscope (Zeiss LSM 510 META with spectral imaging)
- Monochromator-based microscope with dual photodiode detector
- Cell culture facilities
- Transgene mouse facilities
- Cryosection and access to histology facilities
- 96-well Reader for luminescence, fluorescence and photometry (Synergy H4)
- Flow Cytometry (FACSCalibur)
- Radioactivity lab
5 selected publications
1. Birbach A, Eisenbarth D, Kozakowski N, Ladenhauf E, Schmidt-Supprian M, Schmid JA: Persistent inflammation leads to proliferative neoplasia and loss of smooth muscle cells in a prostate tumor model. Neoplasia 2011, 13:692-703.
2. Orel L, Neumeier H, Hochrainer K, Binder BR, Schmid JA: Crosstalk between the NF-κB activating IKK-complex and the CSN signalosome. Journal of cellular and molecular medicine 2010, 14:1555-1568.
3. Schmid JA, Birbach A: IkappaB kinase beta (IKKbeta/IKK2/IKBKB)--a key molecule in signaling to the transcription factor NF-kappaB. Cytokine Growth Factor Rev 2008, 19:157-165.
4. Birbach A, Bailey ST, Ghosh S, Schmid JA: Cytosolic, nuclear and nucleolar localization signals determine subcellular distribution and activity of the NF-kappaB inducing kinase NIK. J Cell Sci 2004, 117:3615-3624.
5. Ebner K, Bandion A, Binder BR, de Martin R, Schmid JA: GMCSF activates NF-kappaB via direct interaction of the GMCSF receptor with IkappaB kinase beta. Blood 2003, 102:192-199.