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Mechanisms of plexin-mediated signalling

Applicant Dr. Jakub Swiercz
Subject Area Pharmacology
Term from 2010 to 2014
Project identifier Deutsche Forschungsgemeinschaft (DFG) - Project number 180324797
 
Final Report Year 2015

Final Report Abstract

In summary, we were able to fulfil the majority of the aims of the project. First, we successfully further characterized Plexin-B1-Met-mediated signalling and identified Grb2 as an important mediator thereof. Furthermore, we could show that a Grb2-dependent signalling mechanism is required for Sema4D-induced, Plexin-B1-mediated regulation of cell migration. These data identify an important new component of plexin-B-mediated signalling and fill the gap between Met-mediated phosphorylation of Plexin-B1 and its ability to deactivate RhoA via p190 RhoGAP. Moreover, on the cellular level, we were able to show that Grb2 is crucial for Plexin-B1-mediated inhibition of cellular motility. In the second part of the project we concentrated on characterization of the Plexin-B1-ErbB-2-mediated signalling pathway. We performed siRNA-based screens and identified the members of the IKK-complex as crucial regulators of Plexin-B1 signalling. On the cellular level, we observed that the Sema4D-dependent, Plexin-B1-mediated migration and dedifferentiation of osteoblasts requires IKK-complex activity, thereby indicating that the significance of the identified IKK signalling mechanism is not only restricted to cancer cells but also extends to other non-malignant cell types. These both findings not only represent a significant advance in our understanding of Plexin-B signalling, but may provide the basis for a future translational effort. Especially the IKK-complex might represent a novel therapeutic target for the treatment of B-plexin-dependent tumours as well as in osteoporosis and other bone diseases. Additionally, we identify over 20,000 phosphorylation sites, of which approximately 1 % is regulated upon TNF-α stimulation. We identify various potential novel IKKβ substrates, including kinases and regulators of cellular trafficking. Moreover, we show that one of the candidates, AEG-1/MTDH/LYRIC, is directly phosphorylated by IKKβ on serine 298. We provide strong evidence that IKKβ-mediated AEG-1 phosphorylation is essential for IκBα degradation as well as NF-κB-dependent gene expression and cell proliferation, which correlate with cancer patient survival in vivo. These findings provide a valuable dataset that can further be used by other scientists exploring IKKβ-mediated signalling. Moreover, we succeeded in describing the way in which AEG-1 regulates IκBα degradation, and we could also show that the levels of AEG-1 phosphorylation correlate with patient survival, thus establishing the possibility of future clinical applications for these findings.

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