Use this URL to cite or link to this record in EThOS:
Title: The modulation of tumour suppressor MST2 and proto-oncogene Raf-1 kinases by the scaffold protein CNK1
Author: Urcia, Roby Joseph
ISNI:       0000 0004 2735 0078
Awarding Body: University of Glasgow
Current Institution: University of Glasgow
Date of Award: 2011
Availability of Full Text:
Access from EThOS:
Access from Institution:
An emerging concept in the regulation of signal transduction specificity is the mediation of scaffold proteins embedded in the circuitry of signalling pathways. The multidomainbased architecture of scaffold proteins facilitates the assembly and modulation of protein complexes to regulate cellular signals to bring about an exacting biological output. The work presented in this thesis aimed to investigate the mechanisms of the protein scaffold CNK1 (connector enhancer of Ras 1) in the pro-apoptotic MST2 pathway and the prooncogenic Raf-1 signalling pathways. Here, by using several molecular, biochemical and cell biology techniques, I demonstrated that CNK1 regulates the interaction of the protooncogene Raf-1 and the tumour suppressor MST2 kinase. Perturbations of CNK1 levels exhibit a biphasic signalling response typical of a scaffold protein. Transient expression of CNK1 upon growth factor withdrawal results in a concentration-dependent increase of the Raf-1/MST2 complex, thus preventing apoptosis, but this complex dissociates at higher expression levels, hence promoting an apoptotic response. Moreover, CNK1 is involved in the regulation of Fas-induced apoptosis via the MST2/RASSF1A pathway by influencing the time-scale kinetics of MST2 docking and release from the Raf- 1/CNK1 complex and its eventual activation. SiRNA-silencing of CNK1 destabilizes the Raf-1 and MST2 interaction, and enhanced MST2/LATS1 interaction that promotes apoptosis. Thus, CNK1 is required for Raf-1 inhibitory function, but is also necessary for MST2-mediated apoptosis. Remarkably, CNK1 selects and switches complex formation of opposing anchored proteins depending on the stimulus. In response to Fas ligand stimulation, MST2 is released, whereas Raf-1 is retained in complex with CNK1. Conversely, CNK1 retains MST2 and whilst releasing Raf-1 from the complex following growth factor treatment. Mapping the multidomain binding sites of CNK1 using peptide array demonstrates specific interaction sites of client protein complexes. Specific CNK1 point mutants were generated, and found to alter wild-type regulation of client protein complexes. Thus, the work described in this thesis may reveal a regulatory crosstalk between the MST2 apoptotic pathway and the Raf-1 proliferative pathway through CNK1 by coordinating assembly of appropriate pathway components to possibly drive discrete stimulus-specific responses.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available
Keywords: QH301 Biology