A comprehensive and functional analysis of B-Raf signalling
Final Report Abstract
The Ras/Raf/MEK/ERK signalling pathway plays a fundamental role in various cellular processes and human disorders. The serine/threonine kinase B-Raf represents the major ERK activator and an important oncoprotein in human cancer. However, little is known about the regulatory network controlling B-Raf. Using biochemical and genetic methods, we established a spatio-temporal catalogue of known and novel proteinprotein interactions and phosphorylation events occurring in the B-Raf signalosome. We also conducted an in-depth analysis of B-Raf homo- and hetero-dimerisation and demonstrated that B-RafV600E, the most common oncogenic B-Raf mutant found in cancer, tends to reside in large dimer interface (DIF) dependent multi-protein complexes, while wildtype B-Raf is predominantly found in smaller ones. We demonstrated that paradoxical ERK activation induced by kinase-dead or inhibitorbound B-Raf requires an intact DIF and that this structural feature plays an essential role in the activation of B-Raf and Raf-1 by oncogenic H- or K-Ras. A key regulatory step in Raf activation is the Ras-induced phosphorylation of the activation loop (TVKS-motif). This project also investigated the mechanisms regulating AL phosphorylation and its importance for B-Raf function under physiological circumstances and for malignant transformation.
Publications
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(2010): Identification and functional characterization of a novel BRAF mutation in pilocytic astrocytoma. International Journal of Cancer, 129, 2297-2303
Eisenhardt, A.E., Olbrich, H., Röring, M., Janzarik, W., Anh, T.N.V., Cin, H., Remke, M., Witt, H., Korshunov, A., Pfister, S.M., Omran, H., and Brummer, T.
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(2011). A novel MCF-10A line allowing conditional oncogene expression in 3D culture. Cell Communication and Signaling, 9:1
Herr, R., Wöhrle, F.U., Danke, C., Berens, C. and Brummer, T.
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(2011). A Novel Recurrent BRAF Fusion Gene Resulting from 7q34 Deletion Comprises an Alternative Mechanism of MAPK Pathway Activation in Pilocytic Astrocytoma. Acta Neuropathologica 121, 763-774
Cin, H., Meyer, C., Herr, R., Janzarik, W.G., Lambert, S., Jones, D.T.W., Jacob, K., Benner, A., Witt, H., Remke, M., Bender, S., Falkenstein, F., Van Anh, T.N., Olbrich, H., von Deimling, A., Pekrun, A., Kulozik, A.E., Gnekow, A.E., Scheurlen, W.,Witt, O., Omran, H., Jabado, N., Collins, V.P., Brummer, T., Marschalek, R., Lichter, P., Korshunov, A. and Pfister, S.M.
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(2011). Strong negative feedback from Erk to Raf confers robustness to MAPK signalling. Molecular Systems Biology, 7:489
Fritsche-Guenther, R., Witzel, F., Sieber, A., Herr, R., Schmidt, N., Braun, S., Brummer, T., Sers, C. and Blüthgen, N.
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(2012). Distinct requirement for an intact dimer interface in wildtype, V600E and kinase-dead B-RAF signaling. The EMBO Journal, 31:2629-47
Röring, M., Herr, R. Fiala, G., Heilmann, K., Braun, S., Eisenhardt, A.E., Halbach, S., Capper, D., von Deimling, A., Schamel, W., Saunders, D.N. and Brummer, T.
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(2012): "Aberrant B-Raf signalling in human cancer – 10 years from bench to bedside". Critical Reviews in Oncogenesis, 17(1), 97–121
Röring, M. and Brummer, T.
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(2013). Kidins220/ARMS associates with B-Raf and the TCR promoting sustained Erk signaling in T cells. Journal of Immunology, 190:1927-35
Deswal, S., Meyer, A., Fiala, G., Eisenhardt, A.E., Schmitt, L.C., Salek, M., Brummer, T., Acuto, O. and Schamel, W.W.A.
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(2013). PKD controls mitotic Golgi complex fragmentation through a Raf-MEK1 pathway. Molecular Biology of the Cell, 24 :222-33
Kienzle, C., Eisler, S.A., Villeneuve, J., Brummer, T., Olayioye, M.A. and Hausser, A.
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(2014). A kinase RNAi screen identified the atypical kinase RIOK-1 as a suppressor of the Ras homologue LET-60 in Caenorhabditis elegans. Gene Expression Patterns 15:124-134
Weinberg, F., Schulze, E., Fatouros, C., Schmidt, E, Baumeister, R.* and T. Brummer
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(2014). BRAF inhibitor-associated ERK activation drives development of chronic lymphocytic leukemia. Journal of Clinical Investigation 124(11):5074-84
Yaktapour, N., Meiss, F., Mastroianni, J., Zenz, T., Andrlova, H., Mathew, N.R., Claus, R., Hutter, B., Fröhling, S., Brors, B., Pfeifer, D., Pantic, M., Bartsch, I., Spehl, T.S., Meyer, P.T., Duyster, J., Zirlik, K., Brummer, T. and R. Zeiser
