Medizinische Universität Graz - Research portal
Selected Publication:
Sailer, B.
THE INTERACTION OF -CATENIN WITH AXIN-1 WITHIN WNT-SIGNALLING
[ Diplomarbeit/Master Thesis (UNI) ] Karl-Franzens-Universität Graz; 2021. pp.132.
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- Authors Med Uni Graz:
- Advisor:
- Madl Tobias
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- Abstract:
- The Wnt pathway is one of the most conserved transduction pathways in mammalian cells; it serves as a key trigger for manifold regulations involving embryogenesis, tissue homeostasis and stem cell renewal. Seen from a holistic perspective the Wnt signalling pathway can be interpreted as a black box with many complex inputs but only one output: - catenin. All the components of this black box, in particular the destruction complex with all its interactions, seem to be built around this very important regulatory protein. If this machinery is switched off by binding of Wnt- ligands, - catenin is degraded insufficiently, leading to an accumulation within the cell and entering the nucleus for inducing the transcription of Wnt target genes. Not surprisingly, aberrant activation is one of the driving forces of various human cancers. In the last two decades the studies of Wnt- signalling provided some crucial insights with the promise of new strategies in creating new therapeutics of maligne disorders. This thesis takes a closer look into a specific gear of the destruction complex,the interaction of Axin-1 with - catenin. A distinct binding site between Axin-1 and catenin has been reported in the past, but lacking of a precise binding location within the sequence. Several binding analyses and the combination of different biophysical methods like Isothermal Titration-Colorimetry (ITC), Fluorescence Polarization (FP)- analysis and Nuclear Magnetic Resonance (NMR)- studies revealed not only the stoichiomtetrics of the binding in vitro but also the finding of an additional binding site which is lying closely to the first one. Both binding sites of Axin-1, positioned at (i) residue number 380-420 and (ii)460-490, are binding the -helical-structured part of the N- terminal armadillo region of catenin. Although this newly discovered site is a way weaker in terms of the binding affinity, meaning it may play a role as a modulator or regulator within the binding mechanism of the destruction complex. In addition, NMR analysis of post-translational modifications like phosphorylation of Axin-1 revealed that the residues serine 359 and threonines 374, 404 and 481 are phosphorylated by the cells kinases. Working with the armadillo segment of - catenin revealed some unexpected properties; the very N- terminal part not only inducesreversible self- oligomerization but is also suspected to alter or modulate the binding activities within this structured region. With this thesis I demonstrate that both binding sites on Axin-1 are initially involved in the binding with - catenin, whereas the first bindingsite features a supportive function upon the binding. Glykogen Synthase-Kinase-3 replaces - catenin at the location of the first binding site, followed by phosphorylation of several serins and threonines, which seems to be prerequisite for interaction of the Axin-1 complex with lipoprotein receptor-related protein 5/6. This thesis puts Axin-1 -catenin interaction in context with the complexity of the destruction complex in Wnt-signalling by connecting different biophysical techniques to build a framework of the molecular mechanisms, that are relevant in physiological and oncological terms.